|
1
|
D'Orso I. The HIV-1 Transcriptional Program: From Initiation to Elongation Control. J Mol Biol 2025; 437:168690. [PMID: 38936695 PMCID: PMC11994015 DOI: 10.1016/j.jmb.2024.168690] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2024] [Revised: 06/20/2024] [Accepted: 06/21/2024] [Indexed: 06/29/2024]
Abstract
A large body of work in the last four decades has revealed the key pillars of HIV-1 transcription control at the initiation and elongation steps. Here, I provide a recount of this collective knowledge starting with the genomic elements (DNA and nascent TAR RNA stem-loop) and transcription factors (cellular and the viral transactivator Tat), and later transitioning to the assembly and regulation of transcription initiation and elongation complexes, and the role of chromatin structure. Compelling evidence support a core HIV-1 transcriptional program regulated by the sequential and concerted action of cellular transcription factors and Tat to promote initiation and sustain elongation, highlighting the efficiency of a small virus to take over its host to produce the high levels of transcription required for viral replication. I summarize new advances including the use of CRISPR-Cas9, genetic tools for acute factor depletion, and imaging to study transcriptional dynamics, bursting and the progression through the multiple phases of the transcriptional cycle. Finally, I describe current challenges to future major advances and discuss areas that deserve more attention to both bolster our basic knowledge of the core HIV-1 transcriptional program and open up new therapeutic opportunities.
Collapse
Affiliation(s)
- Iván D'Orso
- Department of Microbiology, The University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
| |
Collapse
|
|
2
|
Sharma AL, Tyagi P, Khumallambam M, Tyagi M. Cocaine-Induced DNA-Dependent Protein Kinase Relieves RNAP II Pausing by Promoting TRIM28 Phosphorylation and RNAP II Hyperphosphorylation to Enhance HIV Transcription. Cells 2024; 13:1950. [PMID: 39682697 PMCID: PMC11640508 DOI: 10.3390/cells13231950] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2024] [Revised: 11/15/2024] [Accepted: 11/20/2024] [Indexed: 12/18/2024] Open
Abstract
Drug abuse continues to pose a significant challenge in HIV control efforts. In our investigation, we discovered that cocaine not only upregulates the expression of the DNA-dependent protein kinase (DNA-PK) but also augments DNA-PK activation by enhancing its phosphorylation at S2056. Moreover, DNA-PK phosphorylation triggers the higher localization of the DNA-PK into the nucleus. The finding that cocaine increases the nuclear localization of the DNA-PK provides further support to our observation of enhanced DNA-PK recruitment at the HIV long terminal repeat (LTR) following cocaine exposure. By activating and facilitating the nuclear localization of the DNA-PK, cocaine effectively orchestrates multiple stages of HIV transcription, thereby promoting HIV replication. Additionally, our study demonstrates that the cocaine-induced DNA-PK promotes the hyper-phosphorylation of the RNA polymerase II (RNAP II) carboxyl-terminal domain (CTD) at Ser5 and Ser2 sites, enhancing both the initiation and elongation phases, respectively, of HIV transcription. The cocaine-mediated enhancement of transcriptional initiation is supported by its activation of cyclin-dependent kinase 7 (CDK7). Additionally, the induction of transcriptional elongation is marked by higher LTR recruitment and the increased phosphorylation of CDK9, which indicates the stimulation of positive transcriptional elongation factor b (P-TEFb). We demonstrate for the first time that cocaine, through DNA-PK activation, promotes the specific phosphorylation of TRIM28 at serine 824 (p-TRIM28, S824). This modification converts TRIM28 from a transcriptional inhibitor to a transactivator for HIV transcription. Additionally, we observed that the phosphorylation of TRIM28 (p-TRIM28, S824) promotes the transition from the pausing phase to the elongation phase of HIV transcription, thereby facilitating the production of full-length HIV genomic transcripts. This finding corroborates the previously observed enhanced RNAP II CTD phosphorylation at Ser2, a marker of transcriptional elongation, following cocaine exposure. Accordingly, upon cocaine treatment, we observed the elevated recruitment of p-TRIM28-(S824) at the HIV LTR. Overall, our results unravel the intricate molecular mechanisms underlying cocaine-induced HIV transcription and gene expression. These findings hold promise for the development of highly targeted therapeutics aimed at mitigating the detrimental effects of cocaine in individuals living with HIV.
Collapse
Affiliation(s)
| | | | | | - Mudit Tyagi
- Center for Translational Medicine, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA; (A.L.S.); (P.T.); (M.K.)
| |
Collapse
|
|
3
|
Schemelev AN, Davydenko VS, Ostankova YV, Reingardt DE, Serikova EN, Zueva EB, Totolian AA. Involvement of Human Cellular Proteins and Structures in Realization of the HIV Life Cycle: A Comprehensive Review, 2024. Viruses 2024; 16:1682. [PMID: 39599797 PMCID: PMC11599013 DOI: 10.3390/v16111682] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2024] [Revised: 10/25/2024] [Accepted: 10/26/2024] [Indexed: 11/29/2024] Open
Abstract
Human immunodeficiency virus (HIV) continues to be a global health challenge, with over 38 million people infected by the end of 2022. HIV-1, the predominant strain, primarily targets and depletes CD4+ T cells, leading to immunodeficiency and subsequent vulnerability to opportunistic infections. Despite the progress made in antiretroviral therapy (ART), drug resistance and treatment-related toxicity necessitate novel therapeutic strategies. This review delves into the intricate interplay between HIV-1 and host cellular proteins throughout the viral life cycle, highlighting key host factors that facilitate viral entry, replication, integration, and immune evasion. A focus is placed on actual findings regarding the preintegration complex, nuclear import, and the role of cellular cofactors such as FEZ1, BICD2, and NPC components in viral transport and genome integration. Additionally, the mechanisms of immune evasion via HIV-1 proteins Nef and Vpu, and their interaction with host MHC molecules and interferon signaling pathways, are explored. By examining these host-virus interactions, this review underscores the importance of host-targeted therapies in complementing ART, with a particular emphasis on the potential of genetic research and host protein stability in developing innovative treatments for HIV/AIDS.
Collapse
Affiliation(s)
- Alexandr N. Schemelev
- St. Petersburg Pasteur Institute, St. Petersburg 197101, Russia; (V.S.D.); (Y.V.O.); (D.E.R.); (E.N.S.); (E.B.Z.); (A.A.T.)
| | | | | | | | | | | | | |
Collapse
|
|
4
|
Cisneros WJ, Soliman SHA, Walter M, Simons LM, Cornish D, De Fabritiis S, Halle AW, Kim EY, Wolinsky SM, Lorenzo-Redondo R, Shilatifard A, Hultquist JF. Release of P-TEFb from the Super Elongation Complex promotes HIV-1 latency reversal. PLoS Pathog 2024; 20:e1012083. [PMID: 39259751 PMCID: PMC11419360 DOI: 10.1371/journal.ppat.1012083] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/29/2024] [Revised: 09/23/2024] [Accepted: 08/27/2024] [Indexed: 09/13/2024] Open
Abstract
The persistence of HIV-1 in long-lived latent reservoirs during suppressive antiretroviral therapy (ART) remains one of the principal barriers to a functional cure. Blocks to transcriptional elongation play a central role in maintaining the latent state, and several latency reversal strategies focus on the release of positive transcription elongation factor b (P-TEFb) from sequestration by negative regulatory complexes, such as the 7SK complex and BRD4. Another major cellular reservoir of P-TEFb is in Super Elongation Complexes (SECs), which play broad regulatory roles in host gene expression. Still, it is unknown if the release of P-TEFb from SECs is a viable latency reversal strategy. Here, we demonstrate that the SEC is not required for HIV-1 replication in primary CD4+ T cells and that a small molecular inhibitor of the P-TEFb/SEC interaction (termed KL-2) increases viral transcription. KL-2 acts synergistically with other latency reversing agents (LRAs) to reactivate viral transcription in several cell line models of latency in a manner that is, at least in part, dependent on the viral Tat protein. Finally, we demonstrate that KL-2 enhances viral reactivation in peripheral blood mononuclear cells (PBMCs) from people living with HIV (PLWH) on suppressive ART, most notably in combination with inhibitor of apoptosis protein antagonists (IAPi). Taken together, these results suggest that the release of P-TEFb from cellular SECs may be a novel route for HIV-1 latency reactivation.
Collapse
Affiliation(s)
- William J. Cisneros
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Shimaa H. A. Soliman
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Miriam Walter
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Lacy M. Simons
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Daphne Cornish
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Simone De Fabritiis
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Ariel W. Halle
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Eun-Young Kim
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Steven M. Wolinsky
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Ramon Lorenzo-Redondo
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Ali Shilatifard
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| | - Judd F. Hultquist
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, Illinois, United States of America
| |
Collapse
|
|
5
|
Sharma AL, Tyagi P, Khumallambam M, Tyagi M. Cocaine-induced DNA-PK relieves RNAP II pausing by promoting TRIM28 phosphorylation. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.08.19.608673. [PMID: 39229050 PMCID: PMC11370412 DOI: 10.1101/2024.08.19.608673] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 09/05/2024]
Abstract
Drug abuse continues to pose a significant challenge in HIV control efforts. In our investigation, we discovered that cocaine not only upregulates the expression of DNA-dependent protein kinase (DNA-PK) but also augments DNA-PK activation by enhancing its phosphorylation at S2056. Moreover, DNA-PK phosphorylation triggers the translocation of DNA-PK into the nucleus. The finding that cocaine promotes nuclear translocation of DNA-PK further validates our observation of enhanced DNA-PK recruitment at the HIV long terminal repeat (LTR) following cocaine exposure. By activating and facilitating the nuclear translocation of DNA-PK, cocaine effectively orchestrates multiple stages of HIV transcription, thereby promoting HIV replication. Additionally, our study indicates that cocaine-induced DNA-PK promotes hyper-phosphorylation of RNA polymerase II (RNAP II) carboxyl-terminal domain (CTD) at Ser5 and Ser2 sites, enhancing both initiation and elongation phases, respectively, of HIV transcription. Cocaine's enhancement of transcription initiation and elongation is further supported by its activation of cyclin-dependent kinase 7 (CDK7) and subsequent phosphorylation of CDK9, thereby promoting positive transcriptional elongation factor b (P-TEFb) activity. We demonstrate for the first time that cocaine, through DNA-PK activation, promotes the specific phosphorylation of TRIM28 at Serine 824 (p-TRIM28, S824). This modification converts TRIM28 from a transcriptional inhibitor to a transactivator for HIV transcription. Additionally, we observe that phosphorylation of TRIM28 (p-TRIM28, S824) promotes the transition from the pausing phase to the elongation phase of HIV transcription, thereby facilitating the production of full-length HIV genomic transcripts. This finding corroborates the observed enhanced RNAP II CTD phosphorylation at Ser2, a marker of transcriptional elongation, following cocaine exposure. Accordingly, upon cocaine treatment, we observed elevated recruitment of p-TRIM28-(S824) at the HIV LTR. Overall, our results have unraveled the intricate molecular mechanisms underlying cocaine-induced HIV transcription and gene expression. These findings hold promise for the development of highly targeted therapeutics aimed at mitigating the detrimental effects of cocaine in individuals living with HIV.
Collapse
Affiliation(s)
| | - Priya Tyagi
- Center for Translational Medicine, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA
| | - Meenata Khumallambam
- Center for Translational Medicine, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA
| | - Mudit Tyagi
- Center for Translational Medicine, Thomas Jefferson University, 1020 Locust Street, Philadelphia, PA 19107, USA
| |
Collapse
|
|
6
|
Liu Y, Li Q, Song L, Gong C, Tang S, Budinich KA, Vanderbeck A, Mathias KM, Wertheim GB, Nguyen SC, Outen R, Joyce EF, Maillard I, Wan L. Condensate-Promoting ENL Mutation Drives Tumorigenesis In Vivo Through Dynamic Regulation of Histone Modifications and Gene Expression. Cancer Discov 2024; 14:1522-1546. [PMID: 38655899 PMCID: PMC11294821 DOI: 10.1158/2159-8290.cd-23-0876] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2023] [Revised: 02/21/2024] [Accepted: 04/22/2024] [Indexed: 04/26/2024]
Abstract
Gain-of-function mutations in the histone acetylation "reader" eleven-nineteen-leukemia (ENL), found in acute myeloid leukemia (AML) and Wilms tumor, are known to drive condensate formation and gene activation in cellular systems. However, their role in tumorigenesis remains unclear. Using a conditional knock-in mouse model, we show that mutant ENL perturbs normal hematopoiesis, induces aberrant expansion of myeloid progenitors, and triggers rapid onset of aggressive AML. Mutant ENL alters developmental and inflammatory gene programs in part by remodeling histone modifications. Mutant ENL forms condensates in hematopoietic stem/progenitor cells at key leukemogenic genes, and disrupting condensate formation via mutagenesis impairs its chromatin and oncogenic function. Moreover, treatment with an acetyl-binding inhibitor of the mutant ENL displaces these condensates from target loci, inhibits mutant ENL-induced chromatin changes, and delays AML initiation and progression in vivo. Our study elucidates the function of ENL mutations in chromatin regulation and tumorigenesis and demonstrates the potential of targeting pathogenic condensates in cancer treatment. Significance: A direct link between ENL mutations, condensate formation, and tumorigenesis is lacking. This study elucidates the function and mechanism of ENL mutations in leukemogenesis, establishing these mutations as bona fide oncogenic drivers. Our results also support the role of condensate dysregulation in cancer and reveal strategies to target pathogenic condensates.
Collapse
Affiliation(s)
- Yiman Liu
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Qinglan Li
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Lele Song
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Chujie Gong
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Cell and Molecular Biology Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Sylvia Tang
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Krista A. Budinich
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Cancer Biology Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Ashley Vanderbeck
- VMD-PhD Program, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Immunology Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Kaeli M. Mathias
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Biochemistry and Molecular Biophysics Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Center for Computational and Genomic Medicine, The Children’s Hospital of Philadelphia, Philadelphia, Pennsylvania.
| | - Gerald B. Wertheim
- Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Division of Hematopathology, The Children’s Hospital of Philadelphia, Philadelphia, Pennsylvania.
| | - Son C. Nguyen
- Epigenetics Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Department of Genetics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Riley Outen
- Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Eric F. Joyce
- Epigenetics Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Department of Genetics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Ivan Maillard
- Division of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| | - Liling Wan
- Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Epigenetics Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
- Institute for Regenerative Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
| |
Collapse
|
|
7
|
Zhang Z, Xu J, Liu J, Wang J, Lei L. SEC: A core hub during cell fate alteration. FASEB J 2024; 38:e23680. [PMID: 38758186 DOI: 10.1096/fj.202400514r] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2024] [Revised: 04/18/2024] [Accepted: 05/07/2024] [Indexed: 05/18/2024]
Abstract
Pol II pause release is a rate-limiting step in gene transcription, influencing various cell fate alterations. Numerous proteins orchestrate Pol II pause release, thereby playing pivotal roles in the intricate process of cellular fate modulation. Super elongation complex (SEC), a large assembly comprising diverse protein components, has garnered attention due to its emerging significance in orchestrating physiological and pathological cellular identity changes by regulating the transcription of crucial genes. Consequently, SEC emerges as a noteworthy functional complex capable of modulating cell fate alterations. Therefore, a comprehensive review is warranted to systematically summarize the core roles of SEC in different types of cell fate alterations. This review focuses on elucidating the current understanding of the structural and functional basis of SEC. Additionally, we discuss the intricate regulatory mechanisms governing SEC in various models of cell fate alteration, encompassing both physiological and pathological contexts. Furthermore, leveraging the existing knowledge of SEC, we propose some insightful directions for future research, aiming to enhance our mechanistic and functional comprehension of SEC within the diverse landscape of cell fate alterations.
Collapse
Affiliation(s)
- Zhijing Zhang
- College of Life Science, Northeast Agricultural University, Harbin, Heilongjiang Province, China
- Department of Histology and Embryology, Harbin Medical University, Harbin, Heilongjiang Province, China
| | - Jingyi Xu
- Department of Histology and Embryology, Harbin Medical University, Harbin, Heilongjiang Province, China
| | - Jiqiang Liu
- Department of Histology and Embryology, Harbin Medical University, Harbin, Heilongjiang Province, China
| | - Jiaqiang Wang
- College of Life Science, Northeast Agricultural University, Harbin, Heilongjiang Province, China
| | - Lei Lei
- Department of Histology and Embryology, Harbin Medical University, Harbin, Heilongjiang Province, China
| |
Collapse
|
|
8
|
Cisneros WJ, Walter M, Soliman SH, Simons LM, Cornish D, Halle AW, Kim EY, Wolinsky SM, Shilatifard A, Hultquist JF. Release of P-TEFb from the Super Elongation Complex promotes HIV-1 latency reversal. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.03.01.582881. [PMID: 38464055 PMCID: PMC10925308 DOI: 10.1101/2024.03.01.582881] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 03/12/2024]
Abstract
The persistence of HIV-1 in long-lived latent reservoirs during suppressive antiretroviral therapy (ART) remains one of the principal barriers to a functional cure. Blocks to transcriptional elongation play a central role in maintaining the latent state, and several latency reversal strategies focus on the release of positive transcription elongation factor b (P-TEFb) from sequestration by negative regulatory complexes, such as the 7SK complex and BRD4. Another major cellular reservoir of P-TEFb is in Super Elongation Complexes (SECs), which play broad regulatory roles in host gene expression. Still, it is unknown if the release of P-TEFb from SECs is a viable latency reversal strategy. Here, we demonstrate that the SEC is not required for HIV-1 replication in primary CD4+ T cells and that a small molecular inhibitor of the P-TEFb/SEC interaction (termed KL-2) increases viral transcription. KL-2 acts synergistically with other latency reversing agents (LRAs) to reactivate viral transcription in several cell line models of latency in a manner that is, at least in part, dependent on the viral Tat protein. Finally, we demonstrate that KL-2 enhances viral reactivation in peripheral blood mononuclear cells (PBMCs) from people living with HIV on suppressive ART, most notably in combination with inhibitor of apoptosis protein antagonists (IAPi). Taken together, these results suggest that the release of P-TEFb from cellular SECs may be a novel route for HIV-1 latency reactivation.
Collapse
Affiliation(s)
- William J. Cisneros
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Miriam Walter
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Shimaa H.A. Soliman
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Lacy M. Simons
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Daphne Cornish
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Ariel W. Halle
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Eun-Young Kim
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Steven M. Wolinsky
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Ali Shilatifard
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Judd F. Hultquist
- Division of Infectious Diseases, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
- Center for Pathogen Genomics and Microbial Evolution, Havey Institute for Global Health, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| |
Collapse
|
|
9
|
Long Q, Xiang M, Xiao L, Wang J, Guan X, Liu J, Liao C. The Biological Significance of AFF4: Promoting Transcription Elongation, Osteogenic Differentiation and Tumor Progression. Comb Chem High Throughput Screen 2024; 27:1403-1412. [PMID: 37815186 DOI: 10.2174/0113862073241079230920082056] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2022] [Revised: 06/23/2023] [Accepted: 07/27/2023] [Indexed: 10/11/2023]
Abstract
As a member of the AF4/FMR2 (AFF) family, AFF4 is a scaffold protein in the superelongation complex (SEC). In this mini-view, we discuss the role of AFF4 as a transcription elongation factor that mediates HIV activation and replication and stem cell osteogenic differentiation. AFF4 also promotes the progression of head and neck squamous cell carcinoma, leukemia, breast cancer, bladder cancer and other malignant tumors. The biological function of AFF4 is largely achieved through SEC assembly, regulates SRY-box transcription factor 2 (SOX2), MYC, estrogen receptor alpha (ESR1), inhibitor of differentiation 1 (ID1), c-Jun and noncanonical nuclear factor-κB (NF-κB) transcription and combines with fusion in sarcoma (FUS), unique regulatory cyclins (CycT1), or mixed lineage leukemia (MLL). We explore the prospects of using AFF4 as a therapeutic in Acquired immunodeficiency syndrome (AIDS) and malignant tumors and its potential as a stemness regulator.
Collapse
Affiliation(s)
- Qian Long
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| | - Mingli Xiang
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| | - Linlin Xiao
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| | - Jiajia Wang
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| | - Xiaoyan Guan
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| | - Jianguo Liu
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| | - Chengcheng Liao
- Department of Orthodontics II, Affiliated Stomatological Hospital of Zunyi Medical University, Zunyi, 563000, China
- Oral Disease Research Key Laboratory of Guizhou Tertiary Institution, School of Stomatology, Zunyi Medical University, Zunyi, 563006, China
| |
Collapse
|
|
10
|
Aoi Y, Shilatifard A. Transcriptional elongation control in developmental gene expression, aging, and disease. Mol Cell 2023; 83:3972-3999. [PMID: 37922911 DOI: 10.1016/j.molcel.2023.10.004] [Citation(s) in RCA: 24] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2023] [Revised: 09/23/2023] [Accepted: 10/11/2023] [Indexed: 11/07/2023]
Abstract
The elongation stage of transcription by RNA polymerase II (RNA Pol II) is central to the regulation of gene expression in response to developmental and environmental cues in metazoan. Dysregulated transcriptional elongation has been associated with developmental defects as well as disease and aging processes. Decades of genetic and biochemical studies have painstakingly identified and characterized an ensemble of factors that regulate RNA Pol II elongation. This review summarizes recent findings taking advantage of genetic engineering techniques that probe functions of elongation factors in vivo. We propose a revised model of elongation control in this accelerating field by reconciling contradictory results from the earlier biochemical evidence and the recent in vivo studies. We discuss how elongation factors regulate promoter-proximal RNA Pol II pause release, transcriptional elongation rate and processivity, RNA Pol II stability and RNA processing, and how perturbation of these processes is associated with developmental disorders, neurodegenerative disease, cancer, and aging.
Collapse
Affiliation(s)
- Yuki Aoi
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA
| | - Ali Shilatifard
- Simpson Querrey Institute for Epigenetics, Department of Biochemistry and Molecular Genetics Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA.
| |
Collapse
|
|
11
|
Van Gulck E, Pardons M, Nijs E, Verheyen N, Dockx K, Van Den Eynde C, Battivelli E, Vega J, Florence E, Autran B, Archin NM, Margolis DM, Katlama C, Hamimi C, Van Den Wyngaert I, Eyassu F, Vandekerckhove L, Boden D. A truncated HIV Tat demonstrates potent and specific latency reversal activity. Antimicrob Agents Chemother 2023; 67:e0041723. [PMID: 37874295 PMCID: PMC10649039 DOI: 10.1128/aac.00417-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2023] [Accepted: 08/09/2023] [Indexed: 10/25/2023] Open
Abstract
A major barrier to HIV-1 cure is caused by the pool of latently infected CD4 T-cells that persist under combination antiretroviral therapy (cART). This latent reservoir is capable of producing replication-competent infectious viruses once prolonged suppressive cART is withdrawn. Inducing the reactivation of HIV-1 gene expression in T-cells harboring a latent provirus in people living with HIV-1 under cART may result in depletion of this latent reservoir due to cytopathic effects or immune clearance. Studies have investigated molecules that reactivate HIV-1 gene expression, but to date, no latency reversal agent has been identified to eliminate latently infected cells harboring replication-competent HIV in cART-treated individuals. Stochastic fluctuations in HIV-1 tat gene expression have been described and hypothesized to allow the progression into proviral latency. We hypothesized that exposing latently infected CD4+ T-cells to Tat would result in effective latency reversal. Our results indicate the capacity of a truncated Tat protein and mRNA to reactivate HIV-1 in latently infected T-cells ex vivo to a similar degree as the protein kinase C agonist: phorbol 12-myristate 13-acetate, without T-cell activation or any significant transcriptome perturbation.
Collapse
Affiliation(s)
- Ellen Van Gulck
- Janssen Infectious Diseases, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Marion Pardons
- HIV Cure Research Center, Department of Internal Medicine and Pediatrics, Ghent University Hospital, Ghent University, Ghent, Belgium
| | - Erik Nijs
- Janssen Infectious Diseases, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Nick Verheyen
- Janssen Infectious Diseases, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Koen Dockx
- Janssen Infectious Diseases, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Christel Van Den Eynde
- Janssen Infectious Diseases, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Emilie Battivelli
- Janssen Infectious Diseases, A Division of Janssen Pharmaceutica NV, Brisbane, California, USA
| | - Jerel Vega
- Arcturus Therapeutics, Science Center Drive, San Diego, California, USA
| | | | - Brigitte Autran
- Faculty of Medicine Sorbonne-University, CIMI-Paris, UPMC/Inserm, Paris, France
| | - Nancie M. Archin
- University of North Carolina School of Medicine and UNC, HIV Cure Center, Chapel Hill, North Carolina, USA
| | - David M. Margolis
- University of North Carolina School of Medicine and UNC, HIV Cure Center, Chapel Hill, North Carolina, USA
| | - Christine Katlama
- Department Infectious Diseases, Hospital Pitié Salpetière, Sorbonne-University and IPLESP, Paris, France
| | - Chiraz Hamimi
- Faculty of Medicine Sorbonne-University, CIMI-Paris, UPMC/Inserm, Paris, France
| | - Ilse Van Den Wyngaert
- Discovery Sciences, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Filmon Eyassu
- Discovery Sciences, Janssen Research and Development, A Division of Janssen Pharmaceutica NV, Beerse, Belgium
| | - Linos Vandekerckhove
- HIV Cure Research Center, Department of Internal Medicine and Pediatrics, Ghent University Hospital, Ghent University, Ghent, Belgium
| | - Daniel Boden
- Janssen Infectious Diseases, A Division of Janssen Pharmaceutica NV, Brisbane, California, USA
| |
Collapse
|
|
12
|
Jin J, Bai H, Yan H, Deng T, Li T, Xiao R, Fan L, Bai X, Ning H, Liu Z, Zhang K, Wu X, Liang K, Ma P, Gao X, Hu D. PRMT2 promotes HIV-1 latency by preventing nucleolar exit and phase separation of Tat into the Super Elongation Complex. Nat Commun 2023; 14:7274. [PMID: 37949879 PMCID: PMC10638354 DOI: 10.1038/s41467-023-43060-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2023] [Accepted: 10/30/2023] [Indexed: 11/12/2023] Open
Abstract
The HIV-1 Tat protein hijacks the Super Elongation Complex (SEC) to stimulate viral transcription and replication. However, the mechanisms underlying Tat activation and inactivation, which mediate HIV-1 productive and latent infection, respectively, remain incompletely understood. Here, through a targeted complementary DNA (cDNA) expression screening, we identify PRMT2 as a key suppressor of Tat activation, thus contributing to proviral latency in multiple cell line latency models and in HIV-1-infected patient CD4+ T cells. Our data reveal that the transcriptional activity of Tat is oppositely regulated by NPM1-mediated nucleolar retention and AFF4-induced phase separation in the nucleoplasm. PRMT2 preferentially methylates Tat arginine 52 (R52) to reinforce its nucleolar sequestration while simultaneously counteracting its incorporation into the SEC droplets, thereby leading to its functional inactivation to promote proviral latency. Thus, our studies unveil a central and unappreciated role for Tat methylation by PRMT2 in connecting its subnuclear distribution, liquid droplet formation, and transactivating function, which could be therapeutically targeted to eradicate latent viral reservoirs.
Collapse
Affiliation(s)
- Jiaxing Jin
- National Clinical Research Center for Cancer, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, State Key Laboratory of Experimental Hematology, Key Laboratory of Immune Microenvironment and Disease of Ministry of Education, Department of Cell Biology, School of Basic Medicine, Tianjin Medical University Cancer Institute and Hospital, Tianjin Medical University, 300070, Tianjin, China
| | - Hui Bai
- National Clinical Research Center for Cancer, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, State Key Laboratory of Experimental Hematology, Key Laboratory of Immune Microenvironment and Disease of Ministry of Education, Department of Cell Biology, School of Basic Medicine, Tianjin Medical University Cancer Institute and Hospital, Tianjin Medical University, 300070, Tianjin, China
| | - Han Yan
- National Clinical Research Center for Cancer, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, State Key Laboratory of Experimental Hematology, Key Laboratory of Immune Microenvironment and Disease of Ministry of Education, Department of Cell Biology, School of Basic Medicine, Tianjin Medical University Cancer Institute and Hospital, Tianjin Medical University, 300070, Tianjin, China
| | - Ting Deng
- Key Laboratory of Breast Cancer Prevention and Therapy of Ministry of Education, Key Laboratory of Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital, 300060, Tianjin, China
| | - Tianyu Li
- Department of Pathophysiology, School of Basic Medical Sciences, Wuhan University, 430071, Wuhan, China
| | - Ruijing Xiao
- Department of Pathophysiology, School of Basic Medical Sciences, Wuhan University, 430071, Wuhan, China
| | - Lina Fan
- Department of Infectious Diseases, Tianjin Second People's Hospital, Nankai University, 300192, Tianjin, China
| | - Xue Bai
- The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, School of Basic Medical Sciences, Tianjin Medical University, 300070, Tianjin, China
| | - Hanhan Ning
- National Clinical Research Center for Cancer, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, State Key Laboratory of Experimental Hematology, Key Laboratory of Immune Microenvironment and Disease of Ministry of Education, Department of Cell Biology, School of Basic Medicine, Tianjin Medical University Cancer Institute and Hospital, Tianjin Medical University, 300070, Tianjin, China
| | - Zhe Liu
- The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, School of Basic Medical Sciences, Tianjin Medical University, 300070, Tianjin, China
| | - Kai Zhang
- The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, School of Basic Medical Sciences, Tianjin Medical University, 300070, Tianjin, China
| | - Xudong Wu
- The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, School of Basic Medical Sciences, Tianjin Medical University, 300070, Tianjin, China
| | - Kaiwei Liang
- Department of Pathophysiology, School of Basic Medical Sciences, Wuhan University, 430071, Wuhan, China
| | - Ping Ma
- Department of Infectious Diseases, Tianjin Second People's Hospital, Nankai University, 300192, Tianjin, China.
| | - Xin Gao
- State Key Laboratory of Experimental Hematology, National Clinical Research Center for Blood Diseases, Haihe Laboratory of Cell Ecosystem, Institute of Hematology & Blood Diseases Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, 300020, Tianjin, China.
- Tianjin Institutes of Health Science, 301600, Tianjin, China.
| | - Deqing Hu
- National Clinical Research Center for Cancer, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics, State Key Laboratory of Experimental Hematology, Key Laboratory of Immune Microenvironment and Disease of Ministry of Education, Department of Cell Biology, School of Basic Medicine, Tianjin Medical University Cancer Institute and Hospital, Tianjin Medical University, 300070, Tianjin, China.
| |
Collapse
|
|
13
|
D’Orso I, Forst CV. Mathematical Models of HIV-1 Dynamics, Transcription, and Latency. Viruses 2023; 15:2119. [PMID: 37896896 PMCID: PMC10612035 DOI: 10.3390/v15102119] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2023] [Revised: 10/10/2023] [Accepted: 10/18/2023] [Indexed: 10/29/2023] Open
Abstract
HIV-1 latency is a major barrier to curing infections with antiretroviral therapy and, consequently, to eliminating the disease globally. The establishment, maintenance, and potential clearance of latent infection are complex dynamic processes and can be best described with the help of mathematical models followed by experimental validation. Here, we review the use of viral dynamics models for HIV-1, with a focus on applications to the latent reservoir. Such models have been used to explain the multi-phasic decay of viral load during antiretroviral therapy, the early seeding of the latent reservoir during acute infection and the limited inflow during treatment, the dynamics of viral blips, and the phenomenon of post-treatment control. Finally, we discuss that mathematical models have been used to predict the efficacy of potential HIV-1 cure strategies, such as latency-reversing agents, early treatment initiation, or gene therapies, and to provide guidance for designing trials of these novel interventions.
Collapse
Affiliation(s)
- Iván D’Orso
- Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA;
| | - Christian V. Forst
- Department of Genetics and Genomic Sciences, Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
| |
Collapse
|
|
14
|
Ghosh A, Chakraborty P, Biswas D. Fine tuning of the transcription juggernaut: A sweet and sour saga of acetylation and ubiquitination. BIOCHIMICA ET BIOPHYSICA ACTA. GENE REGULATORY MECHANISMS 2023; 1866:194944. [PMID: 37236503 DOI: 10.1016/j.bbagrm.2023.194944] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/17/2023] [Revised: 04/26/2023] [Accepted: 05/18/2023] [Indexed: 05/28/2023]
Abstract
Among post-translational modifications of proteins, acetylation, phosphorylation, and ubiquitination are most extensively studied over the last several decades. Owing to their different target residues for modifications, cross-talk between phosphorylation with that of acetylation and ubiquitination is relatively less pronounced. However, since canonical acetylation and ubiquitination happen only on the lysine residues, an overlap of the same lysine residue being targeted for both acetylation and ubiquitination happens quite frequently and thus plays key roles in overall functional regulation predominantly through modulation of protein stability. In this review, we discuss the cross-talk of acetylation and ubiquitination in the regulation of protein stability for the functional regulation of cellular processes with an emphasis on transcriptional regulation. Further, we emphasize our understanding of the functional regulation of Super Elongation Complex (SEC)-mediated transcription, through regulation of stabilization by acetylation, deacetylation and ubiquitination and associated enzymes and its implication in human diseases.
Collapse
Affiliation(s)
- Avik Ghosh
- Laboratory of Transcription Biology Molecular Genetics Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 32, India
| | - Poushali Chakraborty
- Laboratory of Transcription Biology Molecular Genetics Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 32, India
| | - Debabrata Biswas
- Laboratory of Transcription Biology Molecular Genetics Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 32, India.
| |
Collapse
|
|
15
|
Kenaston MW, Shah PS. The Archer and the Prey: The Duality of PAF1C in Antiviral Immunity. Viruses 2023; 15:1032. [PMID: 37243120 PMCID: PMC10222983 DOI: 10.3390/v15051032] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2023] [Revised: 04/18/2023] [Accepted: 04/21/2023] [Indexed: 05/28/2023] Open
Abstract
In the ongoing arms race between virus and host, fine-tuned gene expression plays a critical role in antiviral signaling. However, viruses have evolved to disrupt this process and promote their own replication by targeting host restriction factors. Polymerase-associated factor 1 complex (PAF1C) is a key player in this relationship, recruiting other host factors to regulate transcription and modulate innate immune gene expression. Consequently, PAF1C is consistently targeted by a diverse range of viruses, either to suppress its antiviral functions or co-opt them for their own benefit. In this review, we delve into the current mechanisms through which PAF1C restricts viruses by activating interferon and inflammatory responses at the transcriptional level. We also highlight how the ubiquity of these mechanisms makes PAF1C especially vulnerable to viral hijacking and antagonism. Indeed, as often as PAF1C is revealed to be a restriction factor, viruses are found to have targeted the complex in reply.
Collapse
Affiliation(s)
- Matthew W. Kenaston
- Department of Microbiology and Molecular Genetics, University of California, Davis, CA 95616, USA
| | - Priya S. Shah
- Department of Microbiology and Molecular Genetics, University of California, Davis, CA 95616, USA
- Department of Chemical Engineering, University of California, Davis, CA 95616, USA
| |
Collapse
|
|
16
|
Chameettachal A, Mustafa F, Rizvi TA. Understanding Retroviral Life Cycle and its Genomic RNA Packaging. J Mol Biol 2023; 435:167924. [PMID: 36535429 DOI: 10.1016/j.jmb.2022.167924] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2022] [Revised: 12/12/2022] [Accepted: 12/12/2022] [Indexed: 12/23/2022]
Abstract
Members of the family Retroviridae are important animal and human pathogens. Being obligate parasites, their replication involves a series of steps during which the virus hijacks the cellular machinery. Additionally, many of the steps of retrovirus replication are unique among viruses, including reverse transcription, integration, and specific packaging of their genomic RNA (gRNA) as a dimer. Progress in retrovirology has helped identify several molecular mechanisms involved in each of these steps, but many are still unknown or remain controversial. This review summarizes our present understanding of the molecular mechanisms involved in various stages of retrovirus replication. Furthermore, it provides a comprehensive analysis of our current understanding of how different retroviruses package their gRNA into the assembling virions. RNA packaging in retroviruses holds a special interest because of the uniqueness of packaging a dimeric genome. Dimerization and packaging are highly regulated and interlinked events, critical for the virus to decide whether its unspliced RNA will be packaged as a "genome" or translated into proteins. Finally, some of the outstanding areas of exploration in the field of RNA packaging are highlighted, such as the role of epitranscriptomics, heterogeneity of transcript start sites, and the necessity of functional polyA sequences. An in-depth knowledge of mechanisms that interplay between viral and cellular factors during virus replication is critical in understanding not only the virus life cycle, but also its pathogenesis, and development of new antiretroviral compounds, vaccines, as well as retroviral-based vectors for human gene therapy.
Collapse
Affiliation(s)
- Akhil Chameettachal
- Department of Microbiology & Immunology, College of Medicine and Health Sciences (CMHS), United Arab Emirates University, Al Ain, United Arab Emirates. https://twitter.com/chameettachal
| | - Farah Mustafa
- Department of Biochemistry, College of Medicine and Health Sciences (CMHS), United Arab Emirates University, Al Ain, United Arab Emirates; Zayed bin Sultan Center for Health Sciences (ZCHS), United Arab Emirates University, Al Ain, United Arab Emirates.
| | - Tahir A Rizvi
- Department of Microbiology & Immunology, College of Medicine and Health Sciences (CMHS), United Arab Emirates University, Al Ain, United Arab Emirates; Zayed bin Sultan Center for Health Sciences (ZCHS), United Arab Emirates University, Al Ain, United Arab Emirates.
| |
Collapse
|
|
17
|
Horvath RM, Brumme ZL, Sadowski I. Inhibition of the TRIM24 bromodomain reactivates latent HIV-1. Sci Rep 2023; 13:556. [PMID: 36631514 PMCID: PMC9832417 DOI: 10.1038/s41598-023-27765-3] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2022] [Accepted: 01/06/2023] [Indexed: 01/12/2023] Open
Abstract
Expression of the HIV-1 genome by RNA Polymerase II is regulated at multiple steps, as are most cellular genes, including recruitment of general transcription factors and control of transcriptional elongation from the core promoter. We recently discovered that tripartite motif protein TRIM24 is recruited to the HIV-1 Long Terminal Repeat (LTR) by interaction with TFII-I and causes transcriptional elongation by stimulating association of PTEF-b/ CDK9. Because TRIM24 is required for stimulation of transcription from the HIV-1 LTR, we were surprised to find that IACS-9571, a specific inhibitor of the TRIM24 C-terminal bromodomain, induces HIV-1 provirus expression in otherwise untreated cells. IACS-9571 reactivates HIV-1 in T cell lines bearing multiple different provirus models of HIV-1 latency. Additionally, treatment with this TRIM24 bromodomain inhibitor encourages productive HIV-1 expression in newly infected cells and inhibits formation of immediate latent transcriptionally repressed provirus. IACS-9571 synergizes with PMA, ionomycin, TNF-α and PEP005 to activate HIV-1 expression. Furthermore, co-treatment of CD4 + T cells from individuals with HIV-1 on antiretroviral therapy (ART) with PEP005 and IACS-9571 caused robust provirus expression. Notably, IACS-9571 did not cause global activation of T cells; rather, it inhibited induction of IL2 and CD69 expression in human PBMCs and Jurkat T cells treated with PEP005 or PMA. These observations indicate the TRIM24 bromodomain inhibitor IACS-9571 represents a novel HIV-1 latency reversing agent (LRA), and unlike other compounds with this activity, causes partial suppression of T cell activation while inducing expression of latent provirus.
Collapse
Affiliation(s)
- Riley M Horvath
- Department of Biochemistry and Molecular Biology, Molecular Epigenetics Group, LSI, University of British Columbia, UBC, 2350 Health Sciences Mall, Vancouver, BC, V6T 1Z3, Canada
| | - Zabrina L Brumme
- Faculty of Health Sciences, Simon Fraser University, Burnaby, BC, Canada
- British Columbia Centre for Excellence in HIV/AIDS, Vancouver, BC, Canada
| | - Ivan Sadowski
- Department of Biochemistry and Molecular Biology, Molecular Epigenetics Group, LSI, University of British Columbia, UBC, 2350 Health Sciences Mall, Vancouver, BC, V6T 1Z3, Canada.
| |
Collapse
|
|
18
|
Tsukumo SI, Subramani PG, Seija N, Tabata M, Maekawa Y, Mori Y, Ishifune C, Itoh Y, Ota M, Fujio K, Di Noia JM, Yasutomo K. AFF3, a susceptibility factor for autoimmune diseases, is a molecular facilitator of immunoglobulin class switch recombination. SCIENCE ADVANCES 2022; 8:eabq0008. [PMID: 36001653 PMCID: PMC9401627 DOI: 10.1126/sciadv.abq0008] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/11/2022] [Accepted: 07/08/2022] [Indexed: 06/15/2023]
Abstract
Immunoglobulin class switch recombination (CSR) plays critical roles in controlling infections and inflammatory tissue injuries. Here, we show that AFF3, a candidate gene for both rheumatoid arthritis and type 1 diabetes, is a molecular facilitator of CSR with an isotype preference. Aff3-deficient mice exhibit low serum levels of immunoglobulins, predominantly immunoglobulin G2c (IgG2c) followed by IgG1 and IgG3 but not IgM. Furthermore, Aff3-deficient mice show weak resistance to Plasmodium yoelii infection, confirming that Aff3 modulates immunity to this pathogen. Mechanistically, the AFF3 protein binds to the IgM and IgG1 switch regions via a C-terminal domain, and Aff3 deficiency reduces the binding of AID to the switch regions less efficiently. One AFF3 risk allele for rheumatoid arthritis is associated with high mRNA expression of AFF3, IGHG2, and IGHA2 in human B cells. These findings demonstrate that AFF3 directly regulates CSR by facilitating the recruitment of AID to the switch regions.
Collapse
Affiliation(s)
- Shin-ichi Tsukumo
- Department of Immunology and Parasitology, Graduate School of Medicine, Tokushima University, Tokushima, Japan
- Department of Interdisciplinary Research on Medicine and Photonics, Institute of Post-LED Photonics, Tokushima University, Tokushima, Japan
| | - Poorani Ganesh Subramani
- Institut de Recherches Cliniques de Montréal, Montréal, QC, Canada
- Department of Medicine and Division of Experimental Medicine, McGill University, Montréal, QC, Canada
| | - Noé Seija
- Institut de Recherches Cliniques de Montréal, Montréal, QC, Canada
- Molecular Biology Programs, Department of Medicine, University of Montreal, Montréal, QC, Canada
| | - Mizuho Tabata
- Department of Parasitology and Infectious Diseases, Gifu University Graduate School of Medicine, Gifu, Japan
| | - Yoichi Maekawa
- Department of Parasitology and Infectious Diseases, Gifu University Graduate School of Medicine, Gifu, Japan
| | - Yuya Mori
- Division of Pathogenesis and Disease Regulation, Department of Pathology, Shiga University of Medical Science, Shiga, Japan
| | - Chieko Ishifune
- Department of Immunology and Parasitology, Graduate School of Medicine, Tokushima University, Tokushima, Japan
| | - Yasushi Itoh
- Division of Pathogenesis and Disease Regulation, Department of Pathology, Shiga University of Medical Science, Shiga, Japan
| | - Mineto Ota
- Department of Allergy and Rheumatology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
- Department of Functional Genomics and Immunological Diseases, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Keishi Fujio
- Department of Allergy and Rheumatology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
| | - Javier M. Di Noia
- Institut de Recherches Cliniques de Montréal, Montréal, QC, Canada
- Department of Medicine and Division of Experimental Medicine, McGill University, Montréal, QC, Canada
- Molecular Biology Programs, Department of Medicine, University of Montreal, Montréal, QC, Canada
| | - Koji Yasutomo
- Department of Immunology and Parasitology, Graduate School of Medicine, Tokushima University, Tokushima, Japan
- Department of Interdisciplinary Research on Medicine and Photonics, Institute of Post-LED Photonics, Tokushima University, Tokushima, Japan
- The Research Cluster Program on Immunological Diseases, Tokushima University, Tokushima, Japan
| |
Collapse
|
|
19
|
Colbran LL, Johnson MR, Mathieson I, Capra JA. Tracing the Evolution of Human Gene Regulation and Its Association with Shifts in Environment. Genome Biol Evol 2021; 13:evab237. [PMID: 34718543 PMCID: PMC8576593 DOI: 10.1093/gbe/evab237] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/16/2021] [Indexed: 12/16/2022] Open
Abstract
As humans populated the world, they adapted to many varying environmental factors, including climate, diet, and pathogens. Because many of these adaptations were mediated by multiple noncoding variants with small effects on gene regulation, it has been difficult to link genomic signals of selection to specific genes, and to describe the regulatory response to selection. To overcome this challenge, we adapted PrediXcan, a machine learning method for imputing gene regulation from genotype data, to analyze low-coverage ancient human DNA (aDNA). First, we used simulated genomes to benchmark strategies for adapting PrediXcan to increase robustness to incomplete data. Applying the resulting models to 490 ancient Eurasians, we found that genes with the strongest divergent regulation among ancient populations with hunter-gatherer, pastoralist, and agricultural lifestyles are enriched for metabolic and immune functions. Next, we explored the contribution of divergent gene regulation to two traits with strong evidence of recent adaptation: dietary metabolism and skin pigmentation. We found enrichment for divergent regulation among genes proposed to be involved in diet-related local adaptation, and the predicted effects on regulation often suggest explanations for known signals of selection, for example, at FADS1, GPX1, and LEPR. In contrast, skin pigmentation genes show little regulatory change over a 38,000-year time series of 2,999 ancient Europeans, suggesting that adaptation mainly involved large-effect coding variants. This work demonstrates that combining aDNA with present-day genomes is informative about the biological differences among ancient populations, the role of gene regulation in adaptation, and the relationship between genetic diversity and complex traits.
Collapse
Affiliation(s)
- Laura L Colbran
- Vanderbilt Genetics Institute, Vanderbilt University Medical Center, USA
- Department of Genetics, Perelman School of Medicine, University of Pennsylvania, USA
| | - Maya R Johnson
- School for Science and Math at Vanderbilt, Vanderbilt University, USA
- Department of Computer Science, Bryn Mawr College, Pennsylvania, USA
| | - Iain Mathieson
- Department of Genetics, Perelman School of Medicine, University of Pennsylvania, USA
| | - John A Capra
- Vanderbilt Genetics Institute, Vanderbilt University Medical Center, USA
- Department of Biological Sciences, Vanderbilt University, USA
- Department of Biomedical Informatics, Vanderbilt University, USA
- Bakar Computational Health Sciences Institute, University of California, San Francisco, USA
- Department of Epidemiology and Biostatistics, University of California, San Francisco, USA
| |
Collapse
|
|
20
|
Walch P, Selkrig J, Knodler LA, Rettel M, Stein F, Fernandez K, Viéitez C, Potel CM, Scholzen K, Geyer M, Rottner K, Steele-Mortimer O, Savitski MM, Holden DW, Typas A. Global mapping of Salmonella enterica-host protein-protein interactions during infection. Cell Host Microbe 2021; 29:1316-1332.e12. [PMID: 34237247 PMCID: PMC8561747 DOI: 10.1016/j.chom.2021.06.004] [Citation(s) in RCA: 47] [Impact Index Per Article: 11.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2020] [Revised: 02/24/2021] [Accepted: 05/21/2021] [Indexed: 11/16/2022]
Abstract
Intracellular bacterial pathogens inject effector proteins to hijack host cellular processes and promote their survival and proliferation. To systematically map effector-host protein-protein interactions (PPIs) during infection, we generated a library of 32 Salmonella enterica serovar Typhimurium (STm) strains expressing chromosomally encoded affinity-tagged effectors and quantified PPIs in macrophages and epithelial cells. We identified 446 effector-host PPIs, 25 of which were previously described, and validated 13 by reciprocal co-immunoprecipitation. While effectors converged on the same host cellular processes, most had multiple targets, which often differed between cell types. We demonstrate that SseJ, SseL, and SifA modulate cholesterol accumulation at the Salmonella-containing vacuole (SCV) partially via the cholesterol transporter Niemann-Pick C1 protein. PipB recruits the organelle contact site protein PDZD8 to the SCV, and SteC promotes actin bundling by phosphorylating formin-like proteins. This study provides a method for probing host-pathogen PPIs during infection and a resource for interrogating STm effector mechanisms.
Collapse
Affiliation(s)
- Philipp Walch
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany; Heidelberg University, Faculty of Biosciences, Heidelberg, Germany
| | - Joel Selkrig
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany
| | - Leigh A Knodler
- Paul G. Allen School for Global Health, College of Veterinary Medicine, Washington State University, Pullman, USA; Laboratory of Intracellular Parasites, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, MT, USA
| | - Mandy Rettel
- EMBL, Proteomics Core Facility, Heidelberg, Germany
| | - Frank Stein
- EMBL, Proteomics Core Facility, Heidelberg, Germany
| | - Keith Fernandez
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany
| | - Cristina Viéitez
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany; EMBL European Bioinformatics Institute, (EMBL-EBI), Hinxton, UK
| | - Clément M Potel
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany
| | - Karoline Scholzen
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany
| | - Matthias Geyer
- Institute of Structural Biology, University of Bonn, Bonn, Germany
| | - Klemens Rottner
- Division of Molecular Cell Biology, Zoological Institute, TU Braunschweig, Braunschweig, Germany; Molecular Cell Biology Group, Helmholtz Centre for Infection Research, Braunschweig, Germany
| | - Olivia Steele-Mortimer
- Laboratory of Intracellular Parasites, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, MT, USA
| | - Mikhail M Savitski
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany; EMBL, Proteomics Core Facility, Heidelberg, Germany
| | - David W Holden
- MRC Centre for Molecular Bacteriology and Infection, Imperial College, London, UK
| | - Athanasios Typas
- European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany.
| |
Collapse
|
|
21
|
Wu F, Nie S, Yao Y, Huo T, Li X, Wu X, Zhao J, Lin YL, Zhang Y, Mo Q, Song Y. Small-molecule inhibitor of AF9/ENL-DOT1L/AF4/AFF4 interactions suppresses malignant gene expression and tumor growth. Theranostics 2021; 11:8172-8184. [PMID: 34373735 PMCID: PMC8344022 DOI: 10.7150/thno.56737] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2020] [Accepted: 06/28/2021] [Indexed: 01/22/2023] Open
Abstract
Chromosome translocations involving mixed lineage leukemia (MLL) gene cause acute leukemia with a poor prognosis. MLL is frequently fused with transcription cofactors AF4 (~35%), AF9 (25%) or its paralog ENL (10%). The AHD domain of AF9/ENL binds to AF4, its paralog AFF4, or histone-H3 lysine-79 (H3K79) methyltransferase DOT1L. Formation of AF9/ENL/AF4/AFF4-containing super elongation complexes (SEC) and the catalytic activity of DOT1L are essential for MLL-rearranged leukemia. Protein-protein interactions (PPI) between AF9/ENL and DOT1L/AF4/AFF4 are therefore a potential drug target. Methods: Compound screening followed by medicinal chemistry was used to find inhibitors of such PPIs, which were examined for their biological activities against MLL-rearranged leukemia and other cancer cells. Results: Compound-1 was identified to be a novel small-molecule inhibitor of the AF9/ENL-DOT1L/AF4/AFF4 interaction with IC50s of 0.9-3.5 µM. Pharmacological inhibition of the PPIs significantly reduced SEC and DOT1L-mediated H3K79 methylation in the leukemia cells. Gene profiling shows compound-1 significantly suppressed the gene signatures related to onco-MLL, DOT1L, HoxA9 and Myc. It selectively inhibited proliferation of onco-MLL- or Myc-driven cancer cells and induced cell differentiation and apoptosis. Compound-1 exhibited strong antitumor activity in a mouse model of MLL-rearranged leukemia. Conclusions: The AF9/ENL-DOT1L/AF4/AFF4 interactions are validated to be an anticancer target and compound-1 is a useful in vivo probe for biological studies as well as a pharmacological lead for further drug development.
Collapse
|
|
22
|
Chen Y, Vos SM, Dienemann C, Ninov M, Urlaub H, Cramer P. Allosteric transcription stimulation by RNA polymerase II super elongation complex. Mol Cell 2021; 81:3386-3399.e10. [PMID: 34265249 DOI: 10.1016/j.molcel.2021.06.019] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2021] [Revised: 05/19/2021] [Accepted: 06/17/2021] [Indexed: 01/11/2023]
Abstract
The super elongation complex (SEC) contains the positive transcription elongation factor b (P-TEFb) and the subcomplex ELL2-EAF1, which stimulates RNA polymerase II (RNA Pol II) elongation. Here, we report the cryoelectron microscopy (cryo-EM) structure of ELL2-EAF1 bound to a RNA Pol II elongation complex at 2.8 Å resolution. The ELL2-EAF1 dimerization module directly binds the RNA Pol II lobe domain, explaining how SEC delivers P-TEFb to RNA Pol II. The same site on the lobe also binds the initiation factor TFIIF, consistent with SEC binding only after the transition from transcription initiation to elongation. Structure-guided functional analysis shows that the stimulation of RNA elongation requires the dimerization module and the ELL2 linker that tethers the module to the RNA Pol II protrusion. Our results show that SEC stimulates elongation allosterically and indicate that this stimulation involves stabilization of a closed conformation of the RNA Pol II active center cleft.
Collapse
Affiliation(s)
- Ying Chen
- Max Planck Institute for Biophysical Chemistry, Department of Molecular Biology, Am Fassberg 11, 37077 Göttingen, Germany
| | - Seychelle M Vos
- Max Planck Institute for Biophysical Chemistry, Department of Molecular Biology, Am Fassberg 11, 37077 Göttingen, Germany
| | - Christian Dienemann
- Max Planck Institute for Biophysical Chemistry, Department of Molecular Biology, Am Fassberg 11, 37077 Göttingen, Germany
| | - Momchil Ninov
- Max Planck Institute for Biophysical Chemistry, Bioanalytical Mass Spectrometry, Am Fassberg 11, 37077 Göttingen, Germany; University Medical Center Göttingen, Institute of Clinical Chemistry, Bioanalytics Group, Robert-Koch-Straße 40, 37075 Göttingen, Germany
| | - Henning Urlaub
- Max Planck Institute for Biophysical Chemistry, Bioanalytical Mass Spectrometry, Am Fassberg 11, 37077 Göttingen, Germany; University Medical Center Göttingen, Institute of Clinical Chemistry, Bioanalytics Group, Robert-Koch-Straße 40, 37075 Göttingen, Germany
| | - Patrick Cramer
- Max Planck Institute for Biophysical Chemistry, Department of Molecular Biology, Am Fassberg 11, 37077 Göttingen, Germany.
| |
Collapse
|
|
23
|
Li X, Song Y. Structure, function and inhibition of critical protein-protein interactions involving mixed lineage leukemia 1 and its fusion oncoproteins. J Hematol Oncol 2021; 14:56. [PMID: 33823889 PMCID: PMC8022399 DOI: 10.1186/s13045-021-01057-7] [Citation(s) in RCA: 38] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/19/2021] [Accepted: 03/05/2021] [Indexed: 12/13/2022] Open
Abstract
Mixed lineage leukemia 1 (MLL1, also known as MLL or KMT2A) is an important transcription factor and histone-H3 lysine-4 (H3K4) methyltransferase. It is a master regulator for transcription of important genes (e.g., Hox genes) for embryonic development and hematopoiesis. However, it is largely dispensable in matured cells. Dysregulation of MLL1 leads to overexpression of certain Hox genes and eventually leukemia initiation. Chromosome translocations involving MLL1 cause ~ 75% of acute leukemia in infants and 5–10% in children and adults with a poor prognosis. Targeted therapeutics against oncogenic fusion MLL1 (onco-MLL1) are therefore needed. Onco-MLL1 consists of the N-terminal DNA-interacting domains of MLL1 fused with one of > 70 fusion partners, among which transcription cofactors AF4, AF9 and its paralog ENL, and ELL are the most frequent. Wild-type (WT)- and onco-MLL1 involve numerous protein–protein interactions (PPI), which play critical roles in regulating gene expression in normal physiology and leukemia. Moreover, WT-MLL1 has been found to be essential for MLL1-rearranged (MLL1-r) leukemia. Rigorous studies of such PPIs have been performed and much progress has been achieved in understanding their structures, structure–function relationships and the mechanisms for activating gene transcription as well as leukemic transformation. Inhibition of several critical PPIs by peptides, peptidomimetic or small-molecule compounds has been explored as a therapeutic approach for MLL1-r leukemia. This review summarizes the biological functions, biochemistry, structure and inhibition of the critical PPIs involving MLL1 and its fusion partner proteins. In addition, challenges and perspectives of drug discovery targeting these PPIs for the treatment of MLL1-r leukemia are discussed.
Collapse
Affiliation(s)
- Xin Li
- Department of Pharmacology and Chemical Biology, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX, 77030, USA
| | - Yongcheng Song
- Department of Pharmacology and Chemical Biology, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX, 77030, USA. .,Dan L. Duncan Comprehensive Cancer Center, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX, 77030, USA.
| |
Collapse
|
|
24
|
Dubey RC, Alam NB, Gaur R. miR-150-mediated increase in glucose uptake in HIV-infected cells. J Med Virol 2020; 93:6377-6382. [PMID: 33368410 DOI: 10.1002/jmv.26755] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2020] [Revised: 12/19/2020] [Accepted: 12/21/2020] [Indexed: 12/19/2022]
Abstract
Replication of HIV-1 inside host cells is dependent on both viral and host factors. MicroRNAs are small noncoding RNAs that regulate protein synthesis. MicroRNAs may control viral replication either by directly targeting the viral genome or indirectly through cellular proteins that are required during the viral lifecycle. HIV infection may, in turn, regulate host microRNA expression to facilitate its propagation inside cells. miR-150 has been reported to be an essential factor involved in T-cell activation and may serve as a biomarker for HIV disease progression. The current study provides valuable insights into the role of miR-150 in HIV infection. We quantified miR-150 expression in HIV-infected Jurkat cells and observed a time-dependent increase in the expression of miR-150. In addition, HIV infection led to an enhanced influx of glucose inside the infected cells, which further increased on overexpression of miR-150. The increased uptake of glucose was due to miR-150-mediated increase in expression of glucose transporter-1 (GLUT1). In an attempt to decipher the mechanism, we identified that HIV Tat protein enhanced the expression of miR-150 which then upregulated GLUT1 in HIV-infected cells. In summary, this study sheds light on the role of miR-150 in HIV infection and paves the way for miR-150 as a novel therapeutic target against HIV-1.
Collapse
Affiliation(s)
- Ravi C Dubey
- Faculty of Life Sciences and Biotechnology, South Asian University, New Delhi, India
| | - Nazmir B Alam
- Faculty of Life Sciences and Biotechnology, South Asian University, New Delhi, India
| | - Ritu Gaur
- Faculty of Life Sciences and Biotechnology, South Asian University, New Delhi, India
| |
Collapse
|
|
25
|
Pinto DO, DeMarino C, Vo TT, Cowen M, Kim Y, Pleet ML, Barclay RA, Noren Hooten N, Evans MK, Heredia A, Batrakova EV, Iordanskiy S, Kashanchi F. Low-Level Ionizing Radiation Induces Selective Killing of HIV-1-Infected Cells with Reversal of Cytokine Induction Using mTOR Inhibitors. Viruses 2020; 12:E885. [PMID: 32823598 PMCID: PMC7472203 DOI: 10.3390/v12080885] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2020] [Revised: 08/07/2020] [Accepted: 08/10/2020] [Indexed: 12/12/2022] Open
Abstract
HIV-1 infects 39.5 million people worldwide, and cART is effective in preventing viral spread by reducing HIV-1 plasma viral loads to undetectable levels. However, viral reservoirs persist by mechanisms, including the inhibition of autophagy by HIV-1 proteins (i.e., Nef and Tat). HIV-1 reservoirs can be targeted by the "shock and kill" strategy, which utilizes latency-reversing agents (LRAs) to activate latent proviruses and immunotarget the virus-producing cells. Yet, limitations include reduced LRA permeability across anatomical barriers and immune hyper-activation. Ionizing radiation (IR) induces effective viral activation across anatomical barriers. Like other LRAs, IR may cause inflammation and modulate the secretion of extracellular vesicles (EVs). We and others have shown that cells may secrete cytokines and viral proteins in EVs and, therefore, LRAs may contribute to inflammatory EVs. In the present study, we mitigated the effects of IR-induced inflammatory EVs (i.e., TNF-α), through the use of mTOR inhibitors (mTORi; Rapamycin and INK128). Further, mTORi were found to enhance the selective killing of HIV-1-infected myeloid and T-cell reservoirs at the exclusion of uninfected cells, potentially via inhibition of viral transcription/translation and induction of autophagy. Collectively, the proposed regimen using cART, IR, and mTORi presents a novel approach allowing for the targeting of viral reservoirs, prevention of immune hyper-activation, and selectively killing latently infected HIV-1 cells.
Collapse
Affiliation(s)
- Daniel O. Pinto
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Catherine DeMarino
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Thy T. Vo
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Maria Cowen
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Yuriy Kim
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Michelle L. Pleet
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Robert A. Barclay
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| | - Nicole Noren Hooten
- Laboratory of Epidemiology and Population Science, National Institute on Aging, National Institutes of Health, Baltimore, MD 21224, USA; (N.N.H.); (M.K.E.)
| | - Michele K. Evans
- Laboratory of Epidemiology and Population Science, National Institute on Aging, National Institutes of Health, Baltimore, MD 21224, USA; (N.N.H.); (M.K.E.)
| | - Alonso Heredia
- Institute of Human Virology, University of Maryland School of Medicine, University of Maryland, Baltimore, MD 21201, USA;
| | - Elena V. Batrakova
- Department of Medicine, University of North Carolina HIV Cure Center; University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, NC 27599, USA;
| | - Sergey Iordanskiy
- Department of Pharmacology and Molecular Therapeutics, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA;
| | - Fatah Kashanchi
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA 20110, USA; (D.O.P.); (C.D.); (T.T.V.); (M.C.); (Y.K.); (M.L.P.); (R.A.B.)
| |
Collapse
|
|
26
|
Walton SJ, Wang H, Quintero-Cadena P, Bateman A, Sternberg PW. Caenorhabditis elegans AF4/FMR2 Family Homolog affl-2 Regulates Heat-Shock-Induced Gene Expression. Genetics 2020; 215:1039-1054. [PMID: 32518061 PMCID: PMC7404228 DOI: 10.1534/genetics.120.302923] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2019] [Accepted: 05/27/2020] [Indexed: 02/08/2023] Open
Abstract
To mitigate the deleterious effects of temperature increases on cellular organization and proteotoxicity, organisms have developed mechanisms to respond to heat stress. In eukaryotes, HSF1 is the master regulator of the heat shock transcriptional response, but the heat shock response pathway is not yet fully understood. From a forward genetic screen for suppressors of heat-shock-induced gene expression in Caenorhabditis elegans, we found a new allele of hsf-1 that alters its DNA-binding domain, and we found three additional alleles of sup-45, a previously molecularly uncharacterized genetic locus. We identified sup-45 as one of the two hitherto unknown C. elegans orthologs of the human AF4/FMR2 family proteins, which are involved in regulation of transcriptional elongation rate. We thus renamed sup-45 as affl-2 (AF4/FMR2-Like). Through RNA-seq, we demonstrated that affl-2 mutants are deficient in heat-shock-induced transcription. Additionally, affl-2 mutants have herniated intestines, while worms lacking its sole paralog (affl-1) appear wild type. AFFL-2 is a broadly expressed nuclear protein, and nuclear localization of AFFL-2 is necessary for its role in heat shock response. affl-2 and its paralog are not essential for proper HSF-1 expression and localization after heat shock, which suggests that affl-2 may function downstream of, or parallel to, hsf-1 Our characterization of affl-2 provides insights into the regulation of heat-shock-induced gene expression to protect against heat stress.
Collapse
Affiliation(s)
- Sophie J Walton
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125
| | - Han Wang
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125
| | - Porfirio Quintero-Cadena
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125
| | - Alex Bateman
- European Molecular Biology Laboratory, European Bioinformatics Institute, (EMBL-EBI), Wellcome Genome Campus, Hinxton, Cambridge CB10 1SD, United Kingdom
| | - Paul W Sternberg
- Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, California 91125
| |
Collapse
|
|
27
|
Wu J, Xue Y, Gao X, Zhou Q. Host cell factors stimulate HIV-1 transcription by antagonizing substrate-binding function of Siah1 ubiquitin ligase to stabilize transcription elongation factor ELL2. Nucleic Acids Res 2020; 48:7321-7332. [PMID: 32479599 PMCID: PMC7367184 DOI: 10.1093/nar/gkaa461] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2019] [Revised: 05/12/2020] [Accepted: 05/19/2020] [Indexed: 12/16/2022] Open
Abstract
The Siah1 and Siah2 ubiquitin ligases are implicated in diverse biological processes ranging from cellular stress responses, signaling to transcriptional regulation. A key substrate of Siah1 is ELL2, which undergoes proteolysis upon polyubiquitination. ELL2 stimulates transcriptional elongation and is a subunit of the Super Elongation Complex (SEC) essential for HIV-1 transactivation. Previously, multiple transcriptional and post-translational mechanisms are reported to control Siah's expression and activity. Here we show that the activity of Siah1/2 can also be suppressed by host cell factor 1 (HCF1), and the hitherto poorly characterized HCF2, which themselves are not degraded but can bind and block the substrate-binding domain (SBD) of Siah1/2 to prevent their autoubiquitination and trans-ubiquitination of downstream targets including ELL2. This effect stabilizes ELL2 and enhances the ELL2-SEC formation for robust HIV-1 transactivation. Thus, our study not only identifies HCF1/2 as novel activators of HIV-1 transcription through inhibiting Siah1 to stabilize ELL2, but also reveals the SBD of Siah1/2 as a previously unrecognized new target for HCF1/2 to exert this inhibition.
Collapse
Affiliation(s)
- Jun Wu
- Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA
| | - Yuhua Xue
- School of Pharmaceutical Sciences, Xiamen University, Xiamen, Fujian 361005, China
| | - Xiang Gao
- School of Pharmaceutical Sciences, Xiamen University, Xiamen, Fujian 361005, China
| | - Qiang Zhou
- Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA
| |
Collapse
|
|
28
|
AFF4 regulates osteogenic differentiation of human dental follicle cells. Int J Oral Sci 2020; 12:20. [PMID: 32606293 PMCID: PMC7327054 DOI: 10.1038/s41368-020-0083-9] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2019] [Revised: 02/21/2020] [Accepted: 02/23/2020] [Indexed: 02/05/2023] Open
Abstract
As a member of the AFF (AF4/FMR2) family, AFF4 is a transcription elongation factor that is a component of the super elongation complex. AFF4 serves as a scaffolding protein that connects transcription factors and promotes gene transcription through elongation and chromatin remodelling. Here, we investigated the effect of AFF4 on human dental follicle cells (DFCs) in osteogenic differentiation. In this study, we found that small interfering RNA-mediated depletion of AFF4 resulted in decreased alkaline phosphatase (ALP) activity and impaired mineralization. In addition, the expression of osteogenic-related genes (DLX5, SP7, RUNX2 and BGLAP) was significantly downregulated. In contrast, lentivirus-mediated overexpression of AFF4 significantly enhanced the osteogenic potential of human DFCs. Mechanistically, we found that both the mRNA and protein levels of ALKBH1, a critical regulator of epigenetics, changed in accordance with AFF4 expression levels. Overexpression of ALKBH1 in AFF4-depleted DFCs partially rescued the impairment of osteogenic differentiation. Our data indicated that AFF4 promoted the osteogenic differentiation of DFCs by upregulating the transcription of ALKBH1.
Collapse
|
|
29
|
Chen Y, Wang Y, Lin W, Sheng R, Wu Y, Xu R, Zhou C, Yuan Q. AFF1 inhibits adipogenic differentiation via targeting TGM2 transcription. Cell Prolif 2020; 53:e12831. [PMID: 32441391 PMCID: PMC7309944 DOI: 10.1111/cpr.12831] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2020] [Revised: 04/16/2020] [Accepted: 04/28/2020] [Indexed: 02/05/2023] Open
Abstract
OBJECTIVES AF4/FMR2 family member 1 (AFF1), known as a central scaffolding protein of super elongation complex (SEC), regulates gene transcription. We previously reported that AFF1 inhibited osteogenic differentiation of human mesenchymal stromal/stem cells (hMSCs). However, its role in adipogenic differentiation has not been elucidated. MATERIALS AND METHODS hMSCs and 3T3-L1 pre-adipocytes were cultured and induced for adipogenic differentiation. Small interfering RNAs (siRNAs) were applied to deplete AFF1 while lentiviruses expressing HA-Aff1 were used for overexpression. Oil Red O staining, triglyceride (TAG) quantification, quantitative real-time PCR (qPCR), Western blot analysis, immunofluorescence staining, RNA sequencing (RNA-seq) analysis and ChIP-qPCR were performed. To evaluate the adipogenesis in vivo, BALB/c nude mice were subcutaneously injected with Aff1-overexpressed 3T3-L1 pre-adipocytes. RESULTS AFF1 depletion leads to an enhanced adipogenesis in both hMSCs and 3T3-L1 pre-adipocytes. Overexpression of Aff1 in 3T3-L1 cells results in the reduction of adipogenic differentiation and less adipose tissue formation in vivo. Mechanistically, AFF1 binds to the promoter region of Tgm2 gene and regulates its transcription. Overexpression of Tgm2 largely rescues adipogenic differentiation of Aff1-deficient cells. CONCLUSIONS Our data indicate that AFF1 inhibits adipogenic differentiation by regulating the transcription of TGM2.
Collapse
Affiliation(s)
- Yaqian Chen
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Yuan Wang
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Weimin Lin
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Rui Sheng
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Yunshu Wu
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Ruoshi Xu
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Chenchen Zhou
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Quan Yuan
- State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| |
Collapse
|
|
30
|
Shukla A, Ramirez NGP, D’Orso I. HIV-1 Proviral Transcription and Latency in the New Era. Viruses 2020; 12:v12050555. [PMID: 32443452 PMCID: PMC7291205 DOI: 10.3390/v12050555] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2020] [Revised: 05/06/2020] [Accepted: 05/12/2020] [Indexed: 12/11/2022] Open
Abstract
Three decades of extensive work in the HIV field have revealed key viral and host cell factors controlling proviral transcription. Various models of transcriptional regulation have emerged based on the collective information from in vitro assays and work in both immortalized and primary cell-based models. Here, we provide a recount of the past and current literature, highlight key regulatory aspects, and further describe potential limitations of previous studies. We particularly delve into critical steps of HIV gene expression including the role of the integration site, nucleosome positioning and epigenomics, and the transition from initiation to pausing and pause release. We also discuss open questions in the field concerning the generality of previous regulatory models to the control of HIV transcription in patients under suppressive therapy, including the role of the heterogeneous integration landscape, clonal expansion, and bottlenecks to eradicate viral persistence. Finally, we propose that building upon previous discoveries and improved or yet-to-be discovered technologies will unravel molecular mechanisms of latency establishment and reactivation in a “new era”.
Collapse
|
|
31
|
Basu S, Nandy A, Biswas D. Keeping RNA polymerase II on the run: Functions of MLL fusion partners in transcriptional regulation. BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2020; 1863:194563. [PMID: 32348849 DOI: 10.1016/j.bbagrm.2020.194563] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/30/2019] [Revised: 01/13/2020] [Accepted: 04/13/2020] [Indexed: 12/21/2022]
Abstract
Since the identification of key MLL fusion partners as transcription elongation factors regulating expression of HOX cluster genes during hematopoiesis, extensive work from the last decade has resulted in significant progress in our overall mechanistic understanding of role of MLL fusion partner proteins in transcriptional regulation of diverse set of genes beyond just the HOX cluster. In this review, we are going to detail overall understanding of role of MLL fusion partner proteins in transcriptional regulation and thus provide mechanistic insights into possible MLL fusion protein-mediated transcriptional misregulation leading to aberrant hematopoiesis and leukemogenesis.
Collapse
Affiliation(s)
- Subham Basu
- Laboratory of Transcription Biology, Molecular Genetics Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 32, India
| | - Arijit Nandy
- Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India
| | - Debabrata Biswas
- Laboratory of Transcription Biology, Molecular Genetics Division, CSIR-Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata 32, India.
| |
Collapse
|
|
32
|
Gao Y, Chen L, Han Y, Wu F, Yang WS, Zhang Z, Huo T, Zhu Y, Yu C, Kim H, Lee M, Tang Z, Phillips K, He B, Jung SY, Song Y, Zhu B, Xu RM, Feng Q. Acetylation of histone H3K27 signals the transcriptional elongation for estrogen receptor alpha. Commun Biol 2020; 3:165. [PMID: 32265480 PMCID: PMC7138820 DOI: 10.1038/s42003-020-0898-0] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2019] [Accepted: 03/18/2020] [Indexed: 12/28/2022] Open
Abstract
As approximately 70% of human breast tumors are estrogen receptor α (ERα)-positive, estrogen and ERα play essential roles in breast cancer development. By interrupting the ERα signaling pathway, endocrine therapy has been proven to be an effective therapeutic strategy. In this study, we identified a mechanism by which Transcription Start Site (TSS)-associated histone H3K27 acetylation signals the Super Elongation Complex (SEC) to regulate transcriptional elongation of the ESR1 (ERα) gene. SEC interacts with H3K27ac on ESR1 TSS through its scaffold protein AFF4. Depletion of AFF4 by siRNA or CRISPR/Cas9 dramatically reduces expression of ESR1 and its target genes, consequently inhibiting breast cancer cell growth. More importantly, a AFF4 mutant which lacks H3K27ac interaction failed to rescue ESR1 gene expression, suggesting H3K27 acetylation at TSS region is a key mark bridging the transition from transcriptional initiation to elongation, and perturbing SEC function can be an alternative strategy for targeting ERα signaling pathway at chromatin level.
Collapse
Affiliation(s)
- Yujing Gao
- Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX, 77204, USA
- Key Laboratory of Fertility Preservation and Maintenance of Ministry of Education, Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Ningxia Medical University, 750004, Yinchuan, China
| | - Lijia Chen
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 100101, Beijing, China
- University of Chinese Academy of Sciences, 100049, Beijing, China
| | - Yali Han
- Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX, 77204, USA
| | - Fangrui Wu
- Department of Pharmacology and Chemical Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Wen-Si Yang
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 100101, Beijing, China
| | - Zheng Zhang
- Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Tong Huo
- Department of Pharmacology and Chemical Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Yingmin Zhu
- Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Chengtai Yu
- Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX, 77204, USA
| | - Hong Kim
- Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX, 77204, USA
| | - Mark Lee
- Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Zhen Tang
- Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Kevin Phillips
- Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Bin He
- Immunology & Transplant Science Center, Department of Surgery and Urology, Houston Methodist Research Institute, Houston, TX, 77030, USA
- Department of Medicine, Weill Cornell Medicine of Cornell University, New York, NY, 10065, USA
| | - Sung Yun Jung
- Department of Biochemistry, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Yongcheng Song
- Department of Pharmacology and Chemical Biology, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Bokai Zhu
- Department of Medicine, University of Pittsburgh, Pittsburgh, PA, 15261, USA
| | - Rui-Ming Xu
- National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 100101, Beijing, China
- University of Chinese Academy of Sciences, 100049, Beijing, China
| | - Qin Feng
- Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston, Houston, TX, 77204, USA.
| |
Collapse
|
|
33
|
Byun S, Han S, Zheng Y, Planelles V, Lee Y. The landscape of alternative splicing in HIV-1 infected CD4 T-cells. BMC Med Genomics 2020; 13:38. [PMID: 32241262 PMCID: PMC7118826 DOI: 10.1186/s12920-020-0680-7] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022] Open
Abstract
Background Elucidating molecular mechanisms that are altered during HIV-1 infection may provide a better understanding of the HIV-1 life cycle and how it interacts with infected T-cells. One such mechanism is alternative splicing (AS), which has been studied for HIV-1 itself, but no systematic analysis has yet been performed on infected T-cells. We hypothesized that AS patterns in infected T-cells may illuminate the molecular mechanisms underlying HIV-1 infection and identify candidate molecular markers for specifically targeting infected T-cells. Methods We downloaded previously published raw RNA-seq data obtained from HIV-1 infected and non-infected T-cells. We estimated percent spliced in (PSI) levels for each AS exon, then identified differential AS events in the infected cells (FDR < 0.05, PSI difference > 0.1). We performed functional gene set enrichment analysis on the genes with differentially expressed AS exons to identify their functional roles. In addition, we used RT-PCR to validate differential alternative splicing events in cyclin T1 (CCNT1) as a case study. Results We identified 427 candidate genes with differentially expressed AS exons in infected T-cells, including 20 genes related to cell surface, 35 to kinases, and 121 to immune-related genes. In addition, protein-protein interaction analysis identified six essential subnetworks related to the viral life cycle, including Transcriptional regulation by TP53, Class I MHC mediated antigen, G2/M transition, and late phase of HIV life cycle. CCNT1 exon 7 was more frequently skipped in infected T-cells, leading to loss of the key Cyclin_N motif and affecting HIV-1 transcriptional elongation. Conclusions Our findings may provide new insight into systemic host AS regulation under HIV-1 infection and may provide useful initial candidates for the discovery of new markers for specifically targeting infected T-cells.
Collapse
Affiliation(s)
- Seyoun Byun
- Department of Biomedical Informatics, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Seonggyun Han
- Department of Biomedical Informatics, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Yue Zheng
- Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Vicente Planelles
- Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA
| | - Younghee Lee
- Department of Biomedical Informatics, University of Utah School of Medicine, Salt Lake City, UT, USA. .,Huntsman Cancer Institute, University of Utah School of Medicine, Salt Lake City, UT, USA.
| |
Collapse
|
|
34
|
Zhang Y, Xiao Q, Wu Z, Xu R, Zou S, Zhou C. AFF4 enhances odontogenic differentiation of human dental pulp cells. Biochem Biophys Res Commun 2020; 525:687-692. [PMID: 32139123 DOI: 10.1016/j.bbrc.2020.02.122] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2020] [Accepted: 02/20/2020] [Indexed: 02/08/2023]
Abstract
AFF4 is a component of super elongation complex (SECs) and functions as a scaffold protein to bridge the transcription elongation factors. It is associated with leukemia, HIV transcription, and head neck cancer. However, its role in odontogenic differentiation of dental pulp cells (DPCs) is unclear. Here, we show the expression of AFF4 is increased during odontogenesis. Depletion of AFF4 in human DPCs leads to a decrease of alkaline phosphatase (ALP) activity, calcium mineralization and odontogenic-related genes expression. On the contrary, Lentivirus-mediated overexpression of AFF4 induces the odontogenic potential of DPCs. Mechanistically, we found AFF4 regulates the transcription of NFIC, a key factor for tooth root formation. Overexpression of NFIC successfully rescues the restricted differentiation of AFF4-depleted cells. Our data demonstrate that AFF4 serves as a previously unknown regulator of odontogenesis.
Collapse
Affiliation(s)
- Yuning Zhang
- State Key Laboratory of Oral Diseases & National Clinical Research Centre for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China
| | - Qingyue Xiao
- State Key Laboratory of Oral Diseases & National Clinical Research Centre for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China
| | - Zuping Wu
- State Key Laboratory of Oral Diseases & National Clinical Research Centre for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China
| | - Ruoshi Xu
- State Key Laboratory of Oral Diseases & National Clinical Research Centre for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China
| | - Shujuan Zou
- State Key Laboratory of Oral Diseases & National Clinical Research Centre for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China; Department of Orthodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China
| | - Chenchen Zhou
- State Key Laboratory of Oral Diseases & National Clinical Research Centre for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China; Department of Pediatric Dentistry, West China Hospital of Stomatology, Sichuan University, Chengdu, 610041, China.
| |
Collapse
|
|
35
|
ELL-associated factors EAF1/2 negatively regulate HIV-1 transcription through inhibition of Super Elongation Complex formation. BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2020; 1863:194508. [PMID: 32087315 DOI: 10.1016/j.bbagrm.2020.194508] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/13/2019] [Revised: 02/16/2020] [Accepted: 02/16/2020] [Indexed: 01/03/2023]
Abstract
The ELL (ELL1 and ELL2)-containing Super Elongation Complex (SEC) is required for efficient HIV-1 transactivation by the viral-encoded Tat protein. EAF1 and EAF2 are ELL-associated factors and considered as positive regulators of ELL. However, their role in HIV-1 transcriptional control is unknown. In this study, we show that EAF1/2 inhibit the SEC-dependent and Tat-activated HIV-1 transcription. EAF1/2 are found to interact with the SEC components in an ELL1/2-dependent manner. Surprisingly, the depletion of EAF1/2 increases the SEC formation and occupancy on the HIV-1 proviral DNA, thereby stimulating Tat transactivation of HIV-1. Although EAF1/2 interact with members of the SEC in a ELL-dependent manner, this interaction competes with the binding of the scaffolding subunit AFF1 with ELL, thus reducing the SEC formation. Together, these data reveal how EAF1/2 regulate the SEC formation to control HIV-1 transcription.
Collapse
|
|
36
|
Chen Y, Cramer P. Structure of the super-elongation complex subunit AFF4 C-terminal homology domain reveals requirements for AFF homo- and heterodimerization. J Biol Chem 2019; 294:10663-10673. [PMID: 31147444 PMCID: PMC6615702 DOI: 10.1074/jbc.ra119.008577] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2019] [Revised: 05/27/2019] [Indexed: 12/12/2022] Open
Abstract
AF4/FMR2 family member 4 (AFF4) is the scaffold protein of the multisubunit super-elongation complex, which plays key roles in the release of RNA polymerase II from promoter-proximal pausing and in the transactivation of HIV-1 transcription. AFF4 consists of an intrinsically disordered N-terminal region that interacts with other super-elongation complex subunits and a C-terminal homology domain (CHD) that is conserved among AF4/FMR2 family proteins, including AFF1, AFF2, AFF3, and AFF4. Here, we solved the X-ray crystal structure of the CHD in human AFF4 (AFF4-CHD) to 2.2 Å resolution and characterized its biochemical properties. The structure disclosed that AFF4-CHD folds into a novel domain that consists of eight helices and is distantly related to tetratrico peptide repeat motifs. Our analyses further revealed that AFF4-CHD mediates the formation of an AFF4 homodimer or an AFF1-AFF4 heterodimer. Results from fluorescence anisotropy experiments suggested that AFF4-CHD interacts with both RNA and DNA in vitro Furthermore, we identified a surface loop region in AFF4-CHD as a substrate for the P-TEFb kinase cyclin-dependent kinase 9, which triggers release of polymerase II from promoter-proximal pausing sites. In conclusion, the AFF-CHD structure and biochemical analyses reported here reveal the molecular basis for the homo- and heterodimerization of AFF proteins and implicate the AFF4-CHD in nucleic acid interactions. The high conservation of the CHD among several other proteins suggests that our results are also relevant for understanding other CHD-containing proteins and their dimerization behavior.
Collapse
Affiliation(s)
- Ying Chen
- From the Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany
| | - Patrick Cramer
- From the Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany
| |
Collapse
|
|
37
|
Gopalan S, Gibbon DM, Banks CA, Zhang Y, Florens LA, Washburn MP, Dabas P, Sharma N, Seidel CW, Conaway RC, Conaway JW. Schizosaccharomyces pombe Pol II transcription elongation factor ELL functions as part of a rudimentary super elongation complex. Nucleic Acids Res 2019; 46:10095-10105. [PMID: 30102332 PMCID: PMC6212713 DOI: 10.1093/nar/gky713] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2018] [Accepted: 07/26/2018] [Indexed: 12/21/2022] Open
Abstract
ELL family transcription factors activate the overall rate of RNA polymerase II (Pol II) transcription elongation by binding directly to Pol II and suppressing its tendency to pause. In metazoa, ELL regulates Pol II transcription elongation as part of a large multisubunit complex referred to as the Super Elongation Complex (SEC), which includes P-TEFb and EAF, AF9 or ENL, and an AFF family protein. Although orthologs of ELL and EAF have been identified in lower eukaryotes including Schizosaccharomyces pombe, it has been unclear whether SEC-like complexes function in lower eukaryotes. In this report, we describe isolation from S. pombe of an ELL-containing complex with features of a rudimentary SEC. This complex includes S. pombe Ell1, Eaf1, and a previously uncharacterized protein we designate Ell1 binding protein 1 (Ebp1), which is distantly related to metazoan AFF family members. Like the metazoan SEC, this S. pombe ELL complex appears to function broadly in Pol II transcription. Interestingly, it appears to have a particularly important role in regulating genes involved in cell separation.
Collapse
Affiliation(s)
- Sneha Gopalan
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA.,The Open University, Milton Keynes, UK
| | - Dana M Gibbon
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA
| | - Charles As Banks
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA
| | - Ying Zhang
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA
| | | | - Michael P Washburn
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA.,Department of Pathology and Laboratory Med icine, University of Kansas Medical Center, Kansas City, KS 66160, USA
| | - Preeti Dabas
- University School of Biotechnology, G.G.S.Indraprastha University, New Delhi 110078, India
| | - Nimisha Sharma
- University School of Biotechnology, G.G.S.Indraprastha University, New Delhi 110078, India
| | | | - Ronald C Conaway
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA.,Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, KS 66160, USA
| | - Joan W Conaway
- Stowers Institute for Medical Research, Kansas City, MO 64110, USA.,Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, KS 66160, USA
| |
Collapse
|
|
38
|
Yu D, Cattoglio C, Xue Y, Zhou Q. A complex between DYRK1A and DCAF7 phosphorylates the C-terminal domain of RNA polymerase II to promote myogenesis. Nucleic Acids Res 2019; 47:4462-4475. [PMID: 30864669 PMCID: PMC6511856 DOI: 10.1093/nar/gkz162] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2018] [Revised: 02/24/2019] [Accepted: 03/02/2019] [Indexed: 12/11/2022] Open
Abstract
The general transcription factor P-TEFb, a master regulator of RNA polymerase (Pol) II elongation, phosphorylates the C-terminal domain (CTD) of Pol II and negative elongation factors to release Pol II from promoter-proximal pausing. We show here that P-TEFb surprisingly inhibits the myoblast differentiation into myotubes, and that P-TEFb and its two positive complexes are eliminated in this process. In contrast, DYRK1A, another CTD kinase known to control transcription of a subset of genes important for development and tissue homeostasis, is found to activate transcription of key myogenic genes. We show that active DYRK1A exists in a complex with the WD40-repeat protein DCAF7 that stabilizes and tethers DYRK1A to Pol II, so that DYRK1A-DCAF7 can co-migrate with and phosphorylate Pol II along the myogenic gene loci. Thus, DCAF7 modulates the kinase signaling output of DYRK1A on Pol II to stimulate myogenic transcription after active P-TEFb function is shut off.
Collapse
Affiliation(s)
- Dan Yu
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
- State Key Laboratory of Cellular Stress Biology, School of Pharmaceutical Sciences, Xiamen University, Xiamen, China
| | - Claudia Cattoglio
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
- Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, CA 94720, USA
| | - Yuhua Xue
- State Key Laboratory of Cellular Stress Biology, School of Pharmaceutical Sciences, Xiamen University, Xiamen, China
| | - Qiang Zhou
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
| |
Collapse
|
|
39
|
Pinto DO, Scott TA, DeMarino C, Pleet ML, Vo TT, Saifuddin M, Kovalskyy D, Erickson J, Cowen M, Barclay RA, Zeng C, Weinberg MS, Kashanchi F. Effect of transcription inhibition and generation of suppressive viral non-coding RNAs. Retrovirology 2019; 16:13. [PMID: 31036006 PMCID: PMC6489247 DOI: 10.1186/s12977-019-0475-0] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2018] [Accepted: 04/20/2019] [Indexed: 01/03/2023] Open
Abstract
Background HIV-1 patients receiving combination antiretroviral therapy (cART) survive infection but require life-long adherence at high expense. In chronic cART-treated patients with undetectable viral titers, cell-associated viral RNA is still detectable, pointing to low-level viral transcriptional leakiness. To date, there are no FDA-approved drugs against HIV-1 transcription. We have previously shown that F07#13, a third generation Tat peptide mimetic with competitive activity against Cdk9/T1-Tat binding sites, inhibits HIV-1 transcription in vitro and in vivo. Results Here, we demonstrate that increasing concentrations of F07#13 (0.01, 0.1, 1 µM) cause a decrease in Tat levels in a dose-dependent manner by inhibiting the Cdk9/T1-Tat complex formation and subsequent ubiquitin-mediated Tat sequestration and degradation. Our data indicate that complexes I and IV contain distinct patterns of ubiquitinated Tat and that transcriptional inhibition induced by F07#13 causes an overall reduction in Tat levels. This reduction may be triggered by F07#13 but ultimately is mediated by TAR-gag viral RNAs that bind suppressive transcription factors (similar to 7SK, NRON, HOTAIR, and Xist lncRNAs) to enhance transcriptional gene silencing and latency. These RNAs complex with PRC2, Sin3A, and Cul4B, resulting in epigenetic modifications. Finally, we observed an F07#13-mediated decrease of viral burden by targeting the R region of the long terminal repeat (HIV-1 promoter region, LTR), promoting both paused polymerases and increased efficiency of CRISPR/Cas9 editing in infected cells. This implies that gene editing may be best performed under a repressed transcriptional state. Conclusions Collectively, our results indicate that F07#13, which can terminate RNA Polymerase II at distinct sites, can generate scaffold RNAs, which may assemble into specific sets of “RNA Machines” that contribute to gene regulation. It remains to be seen whether these effects can also be seen in various clades that have varying promoter strength, mutant LTRs, and in patient samples. Electronic supplementary material The online version of this article (10.1186/s12977-019-0475-0) contains supplementary material, which is available to authorized users.
Collapse
Affiliation(s)
- Daniel O Pinto
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Tristan A Scott
- Center for Gene Therapy, Beckman Research Institute of City of Hope, Duarte, CA, USA
| | - Catherine DeMarino
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Michelle L Pleet
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Thy T Vo
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Mohammed Saifuddin
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Dmytro Kovalskyy
- Protein Engineering Department, Institute of Molecular Biology and Genetics, UAS, Kiev, Ukraine
| | - James Erickson
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Maria Cowen
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Robert A Barclay
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA
| | - Chen Zeng
- Department of Physics, The George Washington University, Washington, DC, USA
| | - Marc S Weinberg
- Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA, USA.,Wits/SA MRC Antiviral Gene Therapy Research Unit, Department of Molecular Medicine and Haematology, University of the Witwatersrand, Johannesburg, South Africa
| | - Fatah Kashanchi
- Laboratory of Molecular Virology, School of Systems Biology, George Mason University, Manassas, VA, USA. .,Laboratory of Molecular Virology, George Mason University, Discovery Hall Room 182, 10900 University Blvd., Manassas, VA, 20110, USA.
| |
Collapse
|
|
40
|
Cat and Mouse: HIV Transcription in Latency, Immune Evasion and Cure/Remission Strategies. Viruses 2019; 11:v11030269. [PMID: 30889861 PMCID: PMC6466452 DOI: 10.3390/v11030269] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/16/2019] [Revised: 03/04/2019] [Accepted: 03/13/2019] [Indexed: 12/13/2022] Open
Abstract
There is broad scientific and societal consensus that finding a cure for HIV infection must be pursued. The major barrier to achieving a cure for HIV/AIDS is the capacity of the HIV virus to avoid both immune surveillance and current antiretroviral therapy (ART) by rapidly establishing latently infected cell populations, termed latent reservoirs. Here, we provide an overview of the rapidly evolving field of HIV cure/remission research, highlighting recent progress and ongoing challenges in the understanding of HIV reservoirs, the role of HIV transcription in latency and immune evasion. We review the major approaches towards a cure that are currently being explored and further argue that small molecules that inhibit HIV transcription, and therefore uncouple HIV gene expression from signals sent by the host immune response, might be a particularly promising approach to attain a cure or remission. We emphasize that a better understanding of the game of "cat and mouse" between the host immune system and the HIV virus is a crucial knowledge gap to be filled in both cure and vaccine research.
Collapse
|
|
41
|
Abstract
BACKGROUND The viral transactivator Tat protein is a key modulator of HIV-1 replication, as it regulates transcriptional elongation from the integrated proviral genome. Tat recruits the human transcription elongation factor b, and other host proteins, such as the super elongation complex, to activate the cellular RNA polymerase II, normally stalled shortly after transcription initiation at the HIV promoter. By means of a complex set of interactions with host cellular factors, Tat determines the fate of viral activity within the infected cell. The virus will either actively replicate to promote dissemination in blood and tissues, or become dormant mostly in memory CD4+ T cells, as part of a small but long-living latent reservoir, the main obstacle for HIV eradication. OBJECTIVE In this review, we summarize recent advances in the understanding of the multi-step mechanism that regulates Tat-mediated HIV-1 transcription and RNA polymerase II release, to promote viral transcription elongation. Early events of the human transcription elongation factor b release from the inhibitory 7SK small nuclear ribonucleoprotein complex and its recruitment to the HIV promoter will be discussed. Specific roles of the super elongation complex subunits during transcription elongation, and insight on recently identified cellular factors and mechanisms regulating HIV latency will be detailed. CONCLUSION Understanding the complexity of HIV transcriptional regulation by host factors may open the door for development of novel strategies to eradicate the resilient latent reservoir.
Collapse
Affiliation(s)
- Guillaume Mousseau
- The Scripps Research Institute, Department of Immunology and Microbiology, 130 Scripps Way, Jupiter, FL 33458. United States
| | - Susana T Valente
- The Scripps Research Institute, Department of Immunology and Microbiology, 130 Scripps Way, Jupiter, FL 33458. United States
| |
Collapse
|
|
42
|
Ne E, Palstra RJ, Mahmoudi T. Transcription: Insights From the HIV-1 Promoter. INTERNATIONAL REVIEW OF CELL AND MOLECULAR BIOLOGY 2018; 335:191-243. [DOI: 10.1016/bs.ircmb.2017.07.011] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
|
|
43
|
Shin Y, Choi BS, Kim KC, Kang C, Kim K, Yoon CH. Development of a dual reporter screening assay for distinguishing the inhibition of HIV Tat-mediated transcription from off-target effects. J Virol Methods 2017; 249:1-9. [PMID: 28807730 DOI: 10.1016/j.jviromet.2017.08.005] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2017] [Revised: 08/07/2017] [Accepted: 08/08/2017] [Indexed: 12/29/2022]
Abstract
Human immunodeficiency virus (HIV) encodes a transcription trans-activator (Tat) with an essential role in the transcriptional elongation of viral RNA based on the viral promoter long terminal repeat (LTR). Tat-mediated transcription is conserved and can be distinguished from host transcription, so it is a therapeutic target for combating HIV replication. Traditional screening assays for Tat-mediated transcriptional inhibitors are based on the biochemical properties of Tat and transactivation-responsive RNA. We developed an inducible system based on two lentiviral expression cassettes for doxycycline (Dox)-inducible Tat and Renilla luciferase (R-Luc) using TZM-bl cells harboring LTR-driven firefly luciferase (F-Luc). The cells simultaneously expressed both Tat-induced F-Luc and R-Luc, so it was possible to recognize off-target effects in the presence of Dox. The system was validated with known inhibitors: CYC202 obtained high sensitivity and specificity, whereas 6Bio and DRB had off-target effects. The MTT-based cytotoxicity test indicated the resistance of the system even at concentrations with off-target effects. The specificity of the system was confirmed using antiretroviral drugs. Our dual reporter system can simply detect Tat inhibitory effects, as well as precisely discriminate between the inhibitory and off-target effects of inhibitors, and may be useful for the development of a therapeutic anti-HIV drug.
Collapse
Affiliation(s)
- YoungHyun Shin
- Division of AIDS, Korea National Institute of Health, Chungbuk, Republic of Korea; Division of Viral Disease Research, Korea National Institute of Health, Chungbuk, Republic of Korea.
| | - Byeong-Sun Choi
- Division of AIDS, Korea National Institute of Health, Chungbuk, Republic of Korea; Division of Viral Disease Research, Korea National Institute of Health, Chungbuk, Republic of Korea.
| | - Kyung-Chang Kim
- Division of AIDS, Korea National Institute of Health, Chungbuk, Republic of Korea; Division of Viral Disease Research, Korea National Institute of Health, Chungbuk, Republic of Korea.
| | - Chun Kang
- Division of AIDS, Korea National Institute of Health, Chungbuk, Republic of Korea; Division of Viral Diseases, Korea National Institute of Health, Chungbuk, Republic of Korea.
| | - Kisoon Kim
- Division of Viral Disease Research, Korea National Institute of Health, Chungbuk, Republic of Korea.
| | - Cheol-Hee Yoon
- Division of AIDS, Korea National Institute of Health, Chungbuk, Republic of Korea; Division of Viral Disease Research, Korea National Institute of Health, Chungbuk, Republic of Korea.
| |
Collapse
|
|
44
|
Liu K, Shen D, Shen J, Gao SM, Li B, Wong C, Feng W, Song Y. The Super Elongation Complex Drives Neural Stem Cell Fate Commitment. Dev Cell 2017; 40:537-551.e6. [PMID: 28350987 DOI: 10.1016/j.devcel.2017.02.022] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2016] [Revised: 01/13/2017] [Accepted: 02/26/2017] [Indexed: 10/19/2022]
Abstract
Asymmetric stem cell division establishes an initial difference between a stem cell and its differentiating sibling, critical for maintaining homeostasis and preventing carcinogenesis. Yet the mechanisms that consolidate and lock in such initial fate bias remain obscure. Here, we use Drosophila neuroblasts to demonstrate that the super elongation complex (SEC) acts as an intrinsic amplifier to drive cell fate commitment. SEC is highly expressed in neuroblasts, where it promotes self-renewal by physically associating with Notch transcription activation complex and enhancing HES (hairy and E(spl)) transcription. HES in turn upregulates SEC activity, forming an unexpected self-reinforcing feedback loop with SEC. SEC inactivation leads to neuroblast loss, whereas its forced activation results in neural progenitor dedifferentiation and tumorigenesis. Our studies unveil an SEC-mediated intracellular amplifier mechanism in ensuring robustness and precision in stem cell fate commitment and provide mechanistic explanation for the highly frequent association of SEC overactivation with human cancers.
Collapse
Affiliation(s)
- Kun Liu
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China; Peking-Tsinghua Center for Life Sciences, Peking University, Beijing 100871, China
| | - Dan Shen
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China
| | - Jingwen Shen
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China
| | - Shihong M Gao
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China
| | - Bo Li
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China
| | - Chouin Wong
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China
| | - Weidong Feng
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China
| | - Yan Song
- Ministry of Education Key Laboratory of Cell Proliferation and Differentiation, School of Life Sciences, Peking University, Beijing 100871, China; Peking-Tsinghua Center for Life Sciences, Peking University, Beijing 100871, China.
| |
Collapse
|
|
45
|
Yang H, Basquin D, Pauli D, Oliver B. Drosophila melanogaster positive transcriptional elongation factors regulate metabolic and sex-biased expression in adults. BMC Genomics 2017; 18:384. [PMID: 28521739 PMCID: PMC5436443 DOI: 10.1186/s12864-017-3755-x] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2017] [Accepted: 05/03/2017] [Indexed: 11/22/2022] Open
Abstract
Background Transcriptional elongation is a generic function, but is also regulated to allow rapid transcription responses. Following relatively long initiation and promoter clearance, RNA polymerase II can pause and then rapidly elongate following recruitment of positive elongation factors. Multiple elongation complexes exist, but the role of specific components in adult Drosophila is underexplored. Results We conducted RNA-seq experiments to analyze the effect of RNAi knockdown of Suppressor of Triplolethal and lilliputian. We similarly analyzed the effect of expressing a dominant negative Cyclin-dependent kinase 9 allele. We observed that almost half of the genes expressed in adults showed reduced expression, supporting a broad role for the three tested genes in steady-state transcript abundance. Expression profiles following lilliputian and Suppressor of Triplolethal RNAi were nearly identical raising the possibility that they are obligatory co-factors. Genes showing reduced expression due to these RNAi treatments were short and enriched for genes encoding metabolic or enzymatic functions. The dominant-negative Cyclin-dependent kinase 9 profiles showed both overlapping and specific differential expression, suggesting involvement in multiple complexes. We also observed hundreds of genes with sex-biased differential expression following treatment. Conclusion Transcriptional profiles suggest that Lilliputian and Suppressor of Triplolethal are obligatory cofactors in the adult and that they can also function with Cyclin-dependent kinase 9 at a subset of loci. Our results suggest that transcriptional elongation control is especially important for rapidly expressed genes to support digestion and metabolism, many of which have sex-biased function. Electronic supplementary material The online version of this article (doi:10.1186/s12864-017-3755-x) contains supplementary material, which is available to authorized users.
Collapse
Affiliation(s)
- Haiwang Yang
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 50 South Drive, Bethesda, MD, 20892, USA.
| | - Denis Basquin
- Department of Genetics & Evolution, Sciences III, University of Geneva, Boulevard d'Yvoy 4, CH 1205, Geneva, Switzerland
| | - Daniel Pauli
- Department of Genetics & Evolution, Sciences III, University of Geneva, Boulevard d'Yvoy 4, CH 1205, Geneva, Switzerland
| | - Brian Oliver
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, 50 South Drive, Bethesda, MD, 20892, USA
| |
Collapse
|
|
46
|
Identification of HIV-1 Tat-Associated Proteins Contributing to HIV-1 Transcription and Latency. Viruses 2017; 9:v9040067. [PMID: 28368303 PMCID: PMC5408673 DOI: 10.3390/v9040067] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2017] [Revised: 03/19/2017] [Accepted: 03/24/2017] [Indexed: 12/31/2022] Open
Abstract
Human immunodeficiency virus type 1 (HIV-1) Tat is a virus-encoded trans-activator that plays a central role in viral transcription. We used our recently developed parallel analysis of in vitro translated open reading frames (ORFs) (PLATO) approach to identify host proteins that associate with HIV-1 Tat. From this proteomic assay, we identify 89 Tat-associated proteins (TAPs). We combine our results with other datasets of Tat or long terminal repeat (LTR)-associated proteins. For some of these proteins (NAT10, TINP1, XRCC5, SIN3A), we confirm their strong association with Tat. These TAPs also suppress Tat-mediated HIV-1 transcription. Removing suppression of HIV-1 transcription benefits the reversal of post-integrated, latent HIV-1 proviruses. We demonstrate that these transcriptionally suppressing TAPs contribute to HIV-1 latency in Jurkat latency (J-LAT) cells. Therefore, our proteomic analysis highlights the previously unappreciated TAPs that play a role in maintaining HIV-1 latency and can be further studied as potential pharmacological targets for the “shock and kill” HIV-1 cure strategy.
Collapse
|
|
47
|
Structural basis for ELL2 and AFF4 activation of HIV-1 proviral transcription. Nat Commun 2017; 8:14076. [PMID: 28134250 PMCID: PMC5290273 DOI: 10.1038/ncomms14076] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2016] [Accepted: 11/28/2016] [Indexed: 02/05/2023] Open
Abstract
The intrinsically disordered scaffold proteins AFF1/4 and the transcription elongation factors ELL1/2 are core components of the super elongation complex required for HIV-1 proviral transcription. Here we report the 2.0-Å resolution crystal structure of the human ELL2 C-terminal domain bound to its 50-residue binding site on AFF4, the ELLBow. The ELL2 domain has the same arch-shaped fold as the tight junction protein occludin. The ELLBow consists of an N-terminal helix followed by an extended hairpin that we refer to as the elbow joint, and occupies most of the concave surface of ELL2. This surface is important for the ability of ELL2 to promote HIV-1 Tat-mediated proviral transcription. The AFF4–ELL2 interface is imperfectly packed, leaving a cavity suggestive of a potential binding site for transcription-promoting small molecules. The host super elongation complex (SEC) is hijacked by HIV-1 for viral transcription. Here the authors present the structure of RNA polymerase elongation factor ELL2 bound to the intrinsically disordered scaffold protein AFF4, identifying an ELL2 surface important for HIV-1 transcription.
Collapse
|
|
48
|
Schulze-Gahmen U, Echeverria I, Stjepanovic G, Bai Y, Lu H, Schneidman-Duhovny D, Doudna JA, Zhou Q, Sali A, Hurley JH. Insights into HIV-1 proviral transcription from integrative structure and dynamics of the Tat:AFF4:P-TEFb:TAR complex. eLife 2016; 5. [PMID: 27731797 PMCID: PMC5072841 DOI: 10.7554/elife.15910] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2016] [Accepted: 10/07/2016] [Indexed: 01/04/2023] Open
Abstract
HIV-1 Tat hijacks the human superelongation complex (SEC) to promote proviral transcription. Here we report the 5.9 Å structure of HIV-1 TAR in complex with HIV-1 Tat and human AFF4, CDK9, and CycT1. The TAR central loop contacts the CycT1 Tat-TAR recognition motif (TRM) and the second Tat Zn2+-binding loop. Hydrogen-deuterium exchange (HDX) shows that AFF4 helix 2 is stabilized in the TAR complex despite not touching the RNA, explaining how it enhances TAR binding to the SEC 50-fold. RNA SHAPE and SAXS data were used to help model the extended (Tat Arginine-Rich Motif) ARM, which enters the TAR major groove between the bulge and the central loop. The structure and functional assays collectively support an integrative structure and a bipartite binding model, wherein the TAR central loop engages the CycT1 TRM and compact core of Tat, while the TAR major groove interacts with the extended Tat ARM. DOI:http://dx.doi.org/10.7554/eLife.15910.001
Collapse
Affiliation(s)
- Ursula Schulze-Gahmen
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States
| | - Ignacia Echeverria
- Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, United States.,Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States.,California Institute of Quantitative Biosciences, University of California San, Francisco, San Francisco, United States
| | - Goran Stjepanovic
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States.,Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, United States
| | - Yun Bai
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States
| | - Huasong Lu
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States
| | - Dina Schneidman-Duhovny
- Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, United States.,Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States.,California Institute of Quantitative Biosciences, University of California San, Francisco, San Francisco, United States
| | - Jennifer A Doudna
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States.,Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, United States.,Howard Hughes Medical Institute, University of California, Berkeley, Berkeley, United States.,Department of Chemistry, University of California, Berkeley, Berkeley, United States
| | - Qiang Zhou
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States
| | - Andrej Sali
- Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, San Francisco, United States.,Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States.,California Institute of Quantitative Biosciences, University of California San, Francisco, San Francisco, United States
| | - James H Hurley
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.,California Institute of Quantitative Biosciences, University of California, Berkeley, Berkeley, United States.,Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, United States
| |
Collapse
|
|
49
|
The Multifaceted Contributions of Chromatin to HIV-1 Integration, Transcription, and Latency. INTERNATIONAL REVIEW OF CELL AND MOLECULAR BIOLOGY 2016; 328:197-252. [PMID: 28069134 DOI: 10.1016/bs.ircmb.2016.08.006] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
Abstract
The capacity of the human immunodeficiency virus (HIV-1) to establish latent infections constitutes a major barrier to the development of a cure for HIV-1. In latent infection, replication competent HIV-1 provirus is integrated within the host genome but remains silent, masking the infected cells from the activity of the host immune response. Despite the progress in elucidating the molecular players that regulate HIV-1 gene expression, the mechanisms driving the establishment and maintenance of latency are still not fully understood. Transcription from the HIV-1 genome occurs in the context of chromatin and is subjected to the same regulatory mechanisms that drive cellular gene expression. Much like in eukaryotic genes, the nucleosomal landscape of the HIV-1 promoter and its position within genomic chromatin are determinants of its transcriptional activity. Understanding the multilayered chromatin-mediated mechanisms that underpin HIV-1 integration and expression is of utmost importance for the development of therapeutic strategies aimed at reducing the pool of latently infected cells. In this review, we discuss the impact of chromatin structure on viral integration, transcriptional regulation and latency, and the host factors that influence HIV-1 replication by regulating chromatin organization. Finally, we describe therapeutic strategies under development to target the chromatin-HIV-1 interplay.
Collapse
|
|
50
|
Zaborowska J, Isa NF, Murphy S. P-TEFb goes viral. Bioessays 2016; 38 Suppl 1:S75-85. [DOI: 10.1002/bies.201670912] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2015] [Revised: 09/23/2015] [Accepted: 09/26/2015] [Indexed: 01/31/2023]
Affiliation(s)
| | - Nur F. Isa
- Sir William Dunn School of Pathology; University of Oxford; Oxford UK
- Department of Biotechnology; Kulliyyah of Science, IIUM; Kuantan Pahang Malaysia
| | - Shona Murphy
- Sir William Dunn School of Pathology; University of Oxford; Oxford UK
| |
Collapse
|