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Yeon KY, Ji S, Cheon HG. Role of activating transcription factor 3 as a mediator of the protective effects of berberine against lipopolysaccharide-stimulated SW982 cells and in rheumatoid arthritis animal models. Toxicol Appl Pharmacol 2025; 497:117279. [PMID: 40010574 DOI: 10.1016/j.taap.2025.117279] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2024] [Revised: 02/20/2025] [Accepted: 02/21/2025] [Indexed: 02/28/2025]
Abstract
This study aimed to explore the protective effects of berberine against rheumatoid arthritis (RA) and clarify the role of activating transcription factor 3 (ATF3) in the mechanism of action of berberine, using a lipopolysaccharide (LPS)-stimulated SW982 human synovial cell line. Berberine treatment resulted in a concentration-dependent reduction in LPS-induced proinflammatory cytokines and matrix metalloproteinases (MMPs) in SW982 cells. These inhibitory effects were associated with increased ATF3 expression, reduced nuclear translocation of nuclear factor-κB (NF-κB), and diminished phosphorylation of mitogen-activated protein kinase (MAPK). In contrast, ATF3 knockdown reversed the suppressive effects of berberine on proinflammatory cytokines and MMP production, leading to enhanced MAPK phosphorylation; however, it had minimal impact on adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. Furthermore, AMPK knockdown negated the protective effects of berberine and reduced ATF3 levels, whereas treatment with 5-aminoimidazole-4-carboxamide ribonucleotide, an AMPK activator, replicated the beneficial effects of berberine. In an in vivo collagen-induced arthritis (CIA) mouse model, intraperitoneal administration of berberine significantly reduced paw edema and arthritis severity, accompanied by ATF3 induction and increased AMPK phosphorylation in the synovial tissue. These findings highlighted the pivotal role of ATF3 in mediating the protective effects of berberine in RA- and LPS-activated synoviocytes, suggesting its potential as a therapeutic agent for RA management.
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Affiliation(s)
- Kwan Yong Yeon
- Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology, Gachon University, Incheon 406-799, Republic of Korea
| | - Seongmi Ji
- Department of Pharmacology, College of Medicine, Gachon University, Incheon 406-799, Republic of Korea
| | - Hyae Gyeong Cheon
- Department of Health Sciences and Technology, Gachon Advanced Institute for Health Sciences and Technology, Gachon University, Incheon 406-799, Republic of Korea; Department of Pharmacology, College of Medicine, Gachon University, Incheon 406-799, Republic of Korea.
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Alcaide Martin A, Bauer R, Führer-Sakel D, Heuer H, Mayerl S. Increased seizure susceptibility in thyroid hormone transporter Mct8/Oatp1c1 knockout mice is associated with altered neurotransmitter systems development. Prog Neurobiol 2025; 247:102731. [PMID: 39986448 DOI: 10.1016/j.pneurobio.2025.102731] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2024] [Revised: 01/14/2025] [Accepted: 02/18/2025] [Indexed: 02/24/2025]
Abstract
Thyroid hormone (TH) transporters such as the monocarboxylate transporter Mct8 and the organic anion transporting protein Oatp1c1 facilitate TH transport into target cells. In humans, inactivating mutations in MCT8 result in Allan-Herndon-Dudley syndrome (AHDS), a severe psychomotor retardation with hallmarks of a central TH deficit and frequently observed seizures of unknown etiology. Here, we aimed to investigate seizure susceptibility in AHDS by using Mct8/Oatp1c1 double-knockout (Dko) mice, a well-established AHDS model. We tested seizure susceptibility using the pilocarpine model and observed a significantly faster occurrence of status epilepticus (SE) and more severe responses to seizure induction in Dko animals. We analyzed neuronal alterations in the hippocampus, an area central in seizure pathology, 12 h after SE by immuno-fluorescence and in situ hybridization (ISH). Dko mice presented increased cFos immunoreactivity, and ectopic expression of somatostatin in CA3 neurons. To unravel underlying mechanisms, we studied neurotransmitter systems in murine hippocampi during development at P12 and in adulthood. Employing immuno-fluorescence, ISH and qPCR analyses, we revealed an abnormal development of the inhibitory GABAergic, excitatory glutamatergic and cholinergic systems in Dko mice. Together, our data point to an altered inhibition/excitation balance in the Dko hippocampus that may explain the increased seizure susceptibility.
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Affiliation(s)
- Andrea Alcaide Martin
- University of Duisburg-Essen, University Hospital Essen, Dept. of Endocrinology, Diabetes and Metabolism, Essen, Germany; Center for Translational Neuro-, and Behavioral Sciences (C-TNBS), Medical Faculty, University of Duisburg-Essen, Essen, Germany
| | - Reinhard Bauer
- Institute of Molecular Cell Biology, Jena University Hospital, Jena, Germany
| | - Dagmar Führer-Sakel
- University of Duisburg-Essen, University Hospital Essen, Dept. of Endocrinology, Diabetes and Metabolism, Essen, Germany; Center for Translational Neuro-, and Behavioral Sciences (C-TNBS), Medical Faculty, University of Duisburg-Essen, Essen, Germany
| | - Heike Heuer
- University of Duisburg-Essen, University Hospital Essen, Dept. of Endocrinology, Diabetes and Metabolism, Essen, Germany; Center for Translational Neuro-, and Behavioral Sciences (C-TNBS), Medical Faculty, University of Duisburg-Essen, Essen, Germany
| | - Steffen Mayerl
- University of Duisburg-Essen, University Hospital Essen, Dept. of Endocrinology, Diabetes and Metabolism, Essen, Germany; Center for Translational Neuro-, and Behavioral Sciences (C-TNBS), Medical Faculty, University of Duisburg-Essen, Essen, Germany.
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Conforti A, Di Girolamo R, Guida M, Alviggi C, Casarini L. Pharmacogenomic of LH and its receptor: are we ready for clinical practice? Reprod Biol Endocrinol 2025; 23:29. [PMID: 40001128 PMCID: PMC11863420 DOI: 10.1186/s12958-025-01359-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/28/2023] [Accepted: 02/05/2025] [Indexed: 02/27/2025] Open
Abstract
Luteinizing hormone (LH) is fundamental to support development and reproduction. It acts through a receptor expressed in the gonads, modulating mitogenic, anti-apoptotic, and steroidogenic signals. LH is also marketed as a drug for controlled ovarian stimulation (COS), where it is administered to women to support the action of follicle-stimulating hormone and can lead to specific responses, depending on the individual genetic background. These concepts underline the relevance of a pharmacogenetic approach to COS, in the attempt to optimize clinical outcomes and avoid adverse events. However, knowledge is currently limited by the paucity of clinical studies. This review aims to provide a comprehensive overview of LH and its receptor activity, starting from the description of their molecular pathways from in vitro studies. Data on LH action from in vivo studies were described, as well as the impact of LH and LH/choriogonadotropin (hCG) receptor genetic variants on folliculogenesis and its association with infertility or polycystic ovarian syndrome. Finally, evidence from clinical studies evaluating genetic polymorphisms in the context of assisted reproductive technology treatments and its implications for a pharmacogenomic approach were discussed.
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Affiliation(s)
- Alessandro Conforti
- Department of Neuroscience, Reproductive Science and Odontostomatology, University of Naples Federico II, Via Sergio Pansini, 5, Napoli, 80131, Italy.
| | | | - Maurizio Guida
- Department of Neuroscience, Reproductive Science and Odontostomatology, University of Naples Federico II, Via Sergio Pansini, 5, Napoli, 80131, Italy
| | - Carlo Alviggi
- Department of Public Health, University of Naples Federico II, Naples, Italy
| | - Livio Casarini
- Unit of Endocrinology, Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, Modena, Italy
- Center for Genomic Research, University of Modena and Reggio Emilia, Modena, Italy
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Yang L, Chen Y, Guo F, Wang B, Ying Z, Kuang Y, Zeng X, Ma L, Yu H, Fu P. Transcription factor ATF3 aggravates kidney fibrosis by maintaining the state of histone H3 lysine 27 acetylation. Chin Med J (Engl) 2025:00029330-990000000-01416. [PMID: 39920894 DOI: 10.1097/cm9.0000000000003425] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/11/2024] [Indexed: 02/10/2025] Open
Abstract
BACKGROUND Chronic kidney disease (CKD) is a global health issue, with renal fibrosis being a common pathway in CKD development. Histone modification plays crucial roles in transcriptional regulation, but their pathological functions and mechanisms in CKD are not well understood. METHODS We utilized chromatin immunoprecipitation with next-generation DNA sequencing (ChIP-seq) and RNA-seq to evaluate the states and functions of H3 lysine 27 acetylation (H3K27ac) and H3 lysine 4 trimethylation (H3K4me3) in kidney of CKD mice. We identified epigenetic factors regulating H3K27ac through motif analysis. Expression of activating transcription factor 3 (ATF3) in CKD mouse models and patients' kidneys was validated via immunofluorescence staining or Western blot. We further generated the Atf3 deficient (Atf3-/-) mice to explore its effect in kidney function and fibrosis. ChIP-seq of H3K27ac from Atf3-/- CKD mice was employed to validate ATF3's regulatory effects. We explored how ATF3 maintains the state of H3K27ac by integrating the data sources from multiple databases. RESULTS The states of H3K27ac and H3K4me3 were changed during CKD, and positively correlated with differential gene expression. ATF3 was highly expressed in kidney of both patients and mice with CKD, and co-localized with H3K27ac in genome, epigenetically regulating H3K27ac state. Atf3 deficient in CKD mice significantly ameliorated kidney dysfunction and fibrotic phenotype, and reduced H3K27ac levels at the ATF3 binding sites. Mechanically, ATF3 may recruit the histone acetyltransferases (HATs) network to maintain the H3K27ac state during CKD. CONCLUSION ATF3 promotes kidney injury and fibrosis in CKD by maintaining the state of H3k27ac via recruiting HATs network.
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Affiliation(s)
- Lina Yang
- Kidney Research Institute, Division of Nephrology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China
| | - Yilong Chen
- Biomedical Big Data Center, West China Hospital, Med-X Center for Informatics, Sichuan University, Chengdu, Sichuan 610041,China
| | - Fan Guo
- Kidney Research Institute, Division of Nephrology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China
| | - Bo Wang
- Kidney Research Institute, Division of Nephrology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China
| | - Zhiye Ying
- Biomedical Big Data Center, West China Hospital, Med-X Center for Informatics, Sichuan University, Chengdu, Sichuan 610041,China
| | - Yalan Kuang
- Biomedical Big Data Center, West China Hospital, Med-X Center for Informatics, Sichuan University, Chengdu, Sichuan 610041,China
| | - Xiaoxi Zeng
- Kidney Research Institute, Division of Nephrology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China
- Biomedical Big Data Center, West China Hospital, Med-X Center for Informatics, Sichuan University, Chengdu, Sichuan 610041,China
| | - Liang Ma
- Kidney Research Institute, Division of Nephrology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China
| | - Haopeng Yu
- Biomedical Big Data Center, West China Hospital, Med-X Center for Informatics, Sichuan University, Chengdu, Sichuan 610041,China
| | - Ping Fu
- Kidney Research Institute, Division of Nephrology, West China Hospital, Sichuan University, Chengdu, Sichuan 610041, China
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Yamauchi I, Sugawa T, Hakata T, Yoshizawa A, Kita T, Kishimoto Y, Kimura S, Sakurai A, Kosugi D, Fujita H, Okamoto K, Ueda Y, Fujii T, Taura D, Sakane Y, Yasoda A, Inagaki N. Transcriptomic landscape of hyperthyroidism in mice overexpressing thyroid-stimulating hormone. iScience 2025; 28:111565. [PMID: 39811643 PMCID: PMC11730581 DOI: 10.1016/j.isci.2024.111565] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2023] [Revised: 07/06/2024] [Accepted: 12/06/2024] [Indexed: 01/16/2025] Open
Abstract
Activation of thyroid-stimulating hormone receptor (TSHR) fundamentally leads to hyperthyroidism. To elucidate TSHR signaling, we conducted transcriptome analyses for hyperthyroid mice that we generated by overexpressing TSH. TSH overexpression drastically changed their thyroid transcriptome. In particular, enrichment analyses identified the cell cycle, phosphatidylinositol 3-kinase/Akt pathway, and Ras-related protein 1 pathway as possibly associated with goiter development. Regarding hyperthyroidism, Slc26a4 was exclusively upregulated with TSH overexpression among genes crucial to thyroid hormone secretion. To verify its significance, we overexpressed TSH in Slc26a4 knockout mice. TSH overexpression caused hyperthyroidism in Slc26a4 knockout mice, equivalent to that in control mice. Thus, we did not observe significant changes in known genes and pathways involved in thyroid hormone secretion with TSH overexpression. Our datasets might include candidate genes that have not yet been identified as regulators of thyroid function. Our transcriptome datasets regarding hyperthyroidism can contribute to future research on TSHR signaling.
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Affiliation(s)
- Ichiro Yamauchi
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Taku Sugawa
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Takuro Hakata
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Akira Yoshizawa
- Department of Otolaryngology-Head and Neck Surgery, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Tomoko Kita
- Department of Otolaryngology-Head and Neck Surgery, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Yo Kishimoto
- Department of Otolaryngology-Head and Neck Surgery, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Sadahito Kimura
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Aya Sakurai
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Daisuke Kosugi
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Haruka Fujita
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Kentaro Okamoto
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Yohei Ueda
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Toshihito Fujii
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Daisuke Taura
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
| | - Yoriko Sakane
- Department of Diabetes, Endocrinology and Nutrition, Graduate School of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8507, Japan
- Sugawa Clinic, Nakagyo-ku, Kyoto 604-8105, Japan
| | - Akihiro Yasoda
- Clinical Research Center, National Hospital Organization Kyoto Medical Center, Fushimi-ku, Kyoto 612-8555, Japan
| | - Nobuya Inagaki
- Medical Research Institute KITANO HOSPITAL, PIIF Tazuke-kofukai, Kita-ku, Osaka 530-8480, Japan
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Marin D, Fernandez GJ, Hernandez JC, Taborda N. A systems biology approach unveils different gene expression control mechanisms governing the immune response genetic program in peripheral blood mononuclear cells exposed to SARS-CoV-2. PLoS One 2024; 19:e0314754. [PMID: 39637135 PMCID: PMC11620636 DOI: 10.1371/journal.pone.0314754] [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: 08/28/2024] [Accepted: 11/16/2024] [Indexed: 12/07/2024] Open
Abstract
COVID-19 and other pandemic viruses continue being important for public health and the global economy. Therefore, it is essential to explore the pathogenesis of COVID-19 more deeply, particularly its association with inflammatory and antiviral processes. In this study, we used the RNA-seq technique to analyze mRNA and non-coding RNA profiles of human peripheral blood mononuclear cells (PBMCs) from healthy individuals after SARS-CoV-2 in vitro exposure, to identify pathways related to immune response and the regulatory post-transcriptional mechanisms triggered that can serve as possible complementary therapeutic targets. Our analyses show that SARS-CoV-2 induced a significant regulation in the expression of 790 genes in PBMCs, of which 733 correspond to mRNAs and 57 to non-coding RNAs (lncRNAs). The immune response, antiviral response, signaling, cell proliferation and metabolism are the main biological processes involved. Among these, the inflammatory response groups the majority of regulated genes with an increase in the expression of chemokines involved in the recruitment of monocytes, neutrophils and T-cells. Additionally, it was observed that exposure to SARS-CoV-2 induces the expression of genes related to the IL-27 pathway but not of IFN-I or IFN-III, indicating the induction of ISGs through this pathway rather than the IFN genes. Moreover, several lncRNA and RNA binding proteins that can act in the cis-regulation of genes of the IL-27 pathway were identified. Our results indicate that SARS-CoV-2 can regulate the expression of multiple genes in PBMCs, mainly related to the inflammatory and antiviral response. Among these, lncRNAs establish an important mechanism in regulating the immune response to the virus. They could contribute to developing severe forms of COVID-19, constituting a possible therapeutic target.
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Affiliation(s)
- Damariz Marin
- GIOM, Facultad de Odontología, Universidad Cooperativa de Colombia, Medellín, Colombia
| | - Geysson Javier Fernandez
- Biología y Control de Enfermedades Infecciosas (BCEI), Universidad de Antioquia- UdeA, Medellín, Colombia
| | - Juan C. Hernandez
- Infettare, Facultad de Medicina, Universidad Cooperativa de Colombia, Medellín, Colombia
- Grupo Inmunovirología, Facultad de Medicina, Universidad de Antioquia- UdeA, Medellín, Colombia
| | - Natalia Taborda
- Corporación Universitaria Remington, Programa de Medicina, Facultad de Ciencias de la Salud, Corporación Universitaria Remington, Medellín, Colombia
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Matsumura S, Fujisawa M, Fujiwara M, Okayama H, Marutani M, Nousou E, Sasaki T, Harada N. CREB coactivator CRTC1 in melanocortin-4 receptor-expressing cells regulate dietary fat intake. FASEB Bioadv 2024; 6:597-611. [PMID: 39650226 PMCID: PMC11618889 DOI: 10.1096/fba.2024-00111] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2024] [Revised: 10/07/2024] [Accepted: 10/14/2024] [Indexed: 12/11/2024] Open
Abstract
Cyclic adenosine monophosphate-response element-binding protein-1-regulated transcription coactivator-1 (CRTC1), a cytoplasmic coactivator that translocates to the nucleus in response to cAMP, is associated with obesity. We previously reported that CRTC1 deficiency in melanocortin-4 receptor (MC4R)-expressing neurons, which regulate appetite and energy metabolism in the brain, causes hyperphagia and obesity under a high-fat diet (HFD). HFD is preferred for mice, and the dietary fat in HFD is the main factor contributing to its palatability. These findings, along with our previous results, suggest that CRTC1 regulates the appetite for dietary fat. Therefore, in this study, we aimed to investigate the dietary fat intake behavior and energy metabolism of MC4R neuron-specific CRTC1 knockout mice fed soybean oil or lard. CRTC1 deficiency increased the intake of soybean oil and significantly increased body weight gain. Furthermore, obesity induced by soybean oil intake was partially due to leptin resistance. No significant changes in soybean oil intake were observed between young CRTC1-deficient and wild-type mice; however, soybean oil intake increased with age. Moreover, lard intake did not significantly affect the body weight. Overall, our findings highlighted the crucial role of CRTC1 in the regulation of spontaneous dietary fat intake. Furthermore, the role of CRTC1 becomes increasingly significant with age.
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Affiliation(s)
| | - Miyu Fujisawa
- Department of NutritionOsaka Metropolitan UniversityHabikino CityOsakaJapan
| | - Mizuki Fujiwara
- Department of NutritionOsaka Metropolitan UniversityHabikino CityOsakaJapan
| | - Houko Okayama
- Department of NutritionOsaka Metropolitan UniversityHabikino CityOsakaJapan
| | - Miona Marutani
- Department of NutritionOsaka Metropolitan UniversityHabikino CityOsakaJapan
| | - Eri Nousou
- Department of NutritionOsaka Metropolitan UniversityHabikino CityOsakaJapan
| | - Tsutomu Sasaki
- Department of Neurology, Graduate School of MedicineOsaka UniversityOsakaJapan
| | - Naoki Harada
- Department of Applied Biological Chemistry, Graduate School of AgricultureOsaka Metropolitan UniversityHabikino CityOsakaJapan
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Pérez-de-Oliveira ME, Wagner VP, Bingle CD, Vargas PA, Bingle L. Disruption of oncogenic pathways in mucoepidermoid carcinoma: CREB inhibitor 666.15 as a potential therapeutic agent. Oral Oncol 2024; 159:107029. [PMID: 39332274 DOI: 10.1016/j.oraloncology.2024.107029] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2024] [Accepted: 09/07/2024] [Indexed: 09/29/2024]
Abstract
OBJECTIVES Mucoepidermoid carcinoma (MEC) is the most common malignant salivary gland tumour with around 50 % of cases carrying the CRTC1-MAML2 translocation. The CREB pathway has been associated with the transforming activity of this translocation. The aim of this study was to determine the effects of CREB inhibition on MEC cell behaviour in vitro. MATERIAL AND METHODS Two translocation-positive (UM-HMC-2 and H292) and one translocation-negative (H253) MEC cell lines were treated with 666.15, a CREB inhibitor. Drug IC50 doses were determined for each cell line. Clonogenic and spheroid assays were used to assess survival, including percentage of cancer stem cells, and transwell and scratch assays evaluated invasive and migratory capacities, respectively. Immunofluorescence staining was used to determine E-cadherin expression. RESULTS CREB inhibition significantly reduced the number of surviving colonies and spheroids and delayed cell invasion in all cell lines, but this was more significant in the fusion positive, UM-HMC-2 cells. The expression of E-cadherin was significantly higher in treated UM-HMC-2 and H292 cells. CONCLUSION CREB inhibition with 666.15 impaired key MEC oncogenic behaviours associated with metastasis and drug resistance, including cell invasion and survival.
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Affiliation(s)
- Maria Eduarda Pérez-de-Oliveira
- Department of Oral Diagnosis, Piracicaba Dental School, Universidade Estadual de Campinas, Piracicaba, São Paulo, Brazil; School of Clinical Dentistry, University of Sheffield, Sheffield, United Kingdom
| | - Vivian Petersen Wagner
- Department of Oral Diagnosis, Piracicaba Dental School, Universidade Estadual de Campinas, Piracicaba, São Paulo, Brazil; School of Clinical Dentistry, University of Sheffield, Sheffield, United Kingdom; Department of Oral Medicine, School of Dentistry, Universidade de São Paulo, São Paulo, São Paulo, Brazil
| | - Colin D Bingle
- Division of Clinical Medicine, School of Medicine and Population Health, University of Sheffield, Sheffield, United Kingdom
| | - Pablo Agustin Vargas
- Department of Oral Diagnosis, Piracicaba Dental School, Universidade Estadual de Campinas, Piracicaba, São Paulo, Brazil
| | - Lynne Bingle
- School of Clinical Dentistry, University of Sheffield, Sheffield, United Kingdom.
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Li R, Pan J, Pan C, Li J, Zhang Z, Shahzad K, Sun Y, Yixi Q, Zhaxi W, Qing H, Song T, Zhao W. Transcriptome analysis of mammary epithelial cell between Sewa sheep and East FriEsian sheep from different localities. BMC Genomics 2024; 25:1038. [PMID: 39501165 PMCID: PMC11539678 DOI: 10.1186/s12864-024-10946-3] [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: 03/30/2024] [Accepted: 10/24/2024] [Indexed: 11/08/2024] Open
Abstract
Mammary epithelial cells, the only milk-producing cell type in the mammary gland, undergo dynamic proliferation and differentiation during pregnancy, culminating in lactation postpartum. The East FriEsian sheep ranks among the world's most prolific dairy breeds, while the Sewa sheep, a unique dual-purpose breed autochthonous to the Qinghai-Tibet Plateau, exhibits significantly lower milk production. Employing tissue culture methods, we successfully established mammary epithelial cell lines from both breeds. Morphological assessment of mammary epithelial cells and immunofluorescence identification of Cytokeratin 7 and Cytokeratin 8 confirmed the epithelial identity of the isolated cells. Subsequent RNA-seq analysis of these in vitro epithelial cell lines revealed 1813 differentially expressed genes (DEGs). Among these, 1108 were significantly up-regulated and 705 were down-regulated in Sewa epithelial sheep cells compared to East FriEsian epithelial cells. KEGG enrichment analysis identified cellular processes, environmental information processing, human diseases, metabolism, and organismal systems as the primary functional categories associated with DEGs. Gene ontology (GO) terms annotation, categorized into molecular function, biological processes, and cellular component, yielded "binding and catalytic activity," "molecular function regulator activity," and "cellular process," "biological regulation," and "regulation of biological process" as the top three terms within each domain, respectively. Clusters of Orthologous Groups of proteins (KOG) classification further revealed that "signal transduction mechanisms" accounted for the largest proportion of DEGs among all KOG categories. Finally, based on these analyses, ATF3 and MPP7 were identified as promising candidate genes for regulating lactation.
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Affiliation(s)
- Rui Li
- College of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, 621010, China
| | - Junru Pan
- College of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, 621010, China
| | - Cheng Pan
- College of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, 621010, China
| | - Jingjing Li
- College of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, 621010, China
| | - Zhenzhen Zhang
- College of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, 621010, China
| | - Khuram Shahzad
- Department of Biosciences, COMSATS University Islamabad, Park Road, Islamabad, 45550, Pakistan
| | - Yu Sun
- College of Animal Science and Technology, Henan Agricultural University, Zhengzhou, Henan, 450002, China
| | - Quzhu Yixi
- Cultural Service Center of Maqian Township, Baingoin County, Nagqu, Xizang, 852599, China
| | - Wangjie Zhaxi
- The Service Station of Agricultural and Animal, Husbandry Technical of Baingoin County, Nagqu, Xizang, 852599, China
| | - Haofeng Qing
- The Service Station of Agricultural and Animal, Husbandry Technical of Baingoin County, Nagqu, Xizang, 852599, China
| | - Tianzeng Song
- Institute of Animal Science, Xizang Academy of Agricultural and Animal Husbandry Science, Lhasa, Xizang, 850009, China.
| | - Wangsheng Zhao
- College of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, 621010, China.
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Omar MH. Disruptions to protein kinase A localization in adrenal pathology. Biochem Soc Trans 2024; 52:2231-2241. [PMID: 39364716 DOI: 10.1042/bst20240444] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2024] [Revised: 08/30/2024] [Accepted: 09/16/2024] [Indexed: 10/05/2024]
Abstract
Cell signaling fidelity requires specificity in protein-protein interactions and precise subcellular localization of signaling molecules. In the case of protein phosphorylation, many kinases and phosphatases exhibit promiscuous substrate pairing and therefore require targeting interactions to modify the appropriate substrates and avoid cross-talk among different pathways. In the past 10 years, researchers have discovered and investigated how loss of specific interactions and subcellular targeting for the protein kinase A catalytic subunit (PKAc) lead to cortisol-producing adenomas and the debilitating stress disorder adrenal Cushing's syndrome. This article reviews classical studies regarding PKA localization in glucocorticoid-producing adrenal cells and synthesizes recent evidence of disrupted PKA localization and selective regulatory interactions in adrenal pathology.
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Affiliation(s)
- Mitchell H Omar
- Department of Biochemistry and Molecular Biology, University of Nevada, Reno, NV 89557, U.S.A
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11
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Liu N, Cui X, Guo T, Wei X, Sun Y, Liu J, Zhang Y, Ma W, Yan W, Chen L. Baicalein Ameliorates Insulin Resistance of HFD/STZ Mice Through Activating PI3K/AKT Signal Pathway of Liver and Skeletal Muscle in a GLP-1R-Dependent Manner. Antioxidants (Basel) 2024; 13:1246. [PMID: 39456499 PMCID: PMC11505556 DOI: 10.3390/antiox13101246] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2024] [Revised: 10/10/2024] [Accepted: 10/10/2024] [Indexed: 10/28/2024] Open
Abstract
Insulin resistance (IR) is the principal pathophysiological change occurring in diabetes mellitus (DM). Baicalein, a bioactive flavonoid primarily extracted from the medicinal plant Scutellaria baicalensis Georgi, has been shown in our previous research to be a potential natural glucagon-like peptide-1 receptor (GLP-1R) agonist. However, the exact therapeutic effect of baicalein on DM and its underlying mechanisms remain elusive. In this study, we investigated the therapeutic effects of baicalein on diabetes and sought to clarify its underlying molecular mechanisms. Our results demonstrated that baicalein improves hyperglycemic, hyperinsulinemic, and glucometabolic disorders in mice with induced diabetes via GLP-1R. This was confirmed by the finding that baicalein's effects on improving IR were largely diminished in mice with whole-body Glp1r ablation. Complementarily, network pharmacology analysis highlighted the pivotal involvement of the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) insulin signaling pathway in the therapeutic actions of baicalein on IR. Our mechanism research significantly confirmed that baicalein mitigates hepatic and muscular IR through the PI3K/AKT signal pathway, both in vitro and in vivo. Furthermore, we demonstrated that baicalein enhances glucose uptake in skeletal muscle cells under IR conditions through the Ca2+/calmodulin-dependent protein kinase II (CaMKII)-adenosine 5'-monophosphate-activated protein kinase (AMPK)-glucose transporter 4 (GLUT4) signaling pathway in a GLP-1R-dependent manner. In conclusion, our findings confirm the therapeutic effects of baicalein on IR and reveal that it improves IR in liver and muscle tissues through the PI3K/AKT insulin signaling pathway in a GLP-1R dependent manner. Moreover, we clarified that baicalein enhances the glucose uptake in skeletal muscle tissue through the Ca2+/CaMKII-AMPK-GLUT4 signal pathway.
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Affiliation(s)
- Na Liu
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Xin Cui
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Tingli Guo
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Xiaotong Wei
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Yuzhuo Sun
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Jieyun Liu
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Yangyang Zhang
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Weina Ma
- School of Pharmacy, Xi’an Jiaotong University, Xi’an 710049, China;
| | - Wenhui Yan
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
| | - Lina Chen
- Department of Pharmacology, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an 710061, China; (N.L.); (X.C.); (T.G.); (X.W.); (Y.S.); (J.L.); (Y.Z.)
- Key Laboratory of Environment and Genes Related to Diseases (Xi’an Jiaotong University), Ministry of Education, Xi’an 710061, China
- Cardiometabolic Innovation Center, Ministry of Education, Xi’an 710061, China
- Department of Endocrinology and Second Department of Geriatrics, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710061, China
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12
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Armando NG, Dos Santos Claro PA, Fuertes M, Arzt E, Silberstein S. Role of canonical and non-canonical cAMP sources in CRHR2α-dependent signaling. PLoS One 2024; 19:e0310699. [PMID: 39356686 PMCID: PMC11446442 DOI: 10.1371/journal.pone.0310699] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2023] [Accepted: 09/05/2024] [Indexed: 10/04/2024] Open
Abstract
Hippocampal neurons exhibit activation of both the conventional transmembrane adenylyl cyclases (tmACs) and the non-canonical soluble adenylyl cyclase (sAC) as sources of cyclic AMP (cAMP). These two cAMP sources play crucial roles in mediating signaling pathways downstream of CRHR1 in neuronal and neuroendocrine contexts. In this study, we investigate the involvement of both cAMP sources in the molecular mechanisms triggered by CRHR2α. Here we provide evidence demonstrating that UCN1 and UCN3 exert a neuritogenic effect on HT22-CRHR2α cells, which is solely dependent on the cAMP pool generated by sAC and PKA activity but independent of ERK1/2 activation. Through the characterization of the effectors implicated in neurite elongation, we found that CREB phosphorylation and c-Fos induction rely on PKA activity and ERK1/2 phosphorylation, underscoring the critical role of signaling pathway regulation. These findings strengthen the concept that localized cAMP microdomains actively participate in the regulation of these signaling processes.
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Affiliation(s)
- Natalia G Armando
- Instituto de Investigación en Biomedicina de Buenos Aires (IBioBA), CONICET, Partner Institute of the Max Planck Society, Buenos Aires, Argentina
- Departamento de Ciencia y Tecnología, Universidad Nacional de Quilmes, Buenos Aires, Argentina
| | - Paula A Dos Santos Claro
- Instituto de Investigación en Biomedicina de Buenos Aires (IBioBA), CONICET, Partner Institute of the Max Planck Society, Buenos Aires, Argentina
| | - Mariana Fuertes
- Instituto de Investigación en Biomedicina de Buenos Aires (IBioBA), CONICET, Partner Institute of the Max Planck Society, Buenos Aires, Argentina
- Departamento de Fisiología y Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Eduardo Arzt
- Instituto de Investigación en Biomedicina de Buenos Aires (IBioBA), CONICET, Partner Institute of the Max Planck Society, Buenos Aires, Argentina
- Departamento de Fisiología y Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Susana Silberstein
- Instituto de Investigación en Biomedicina de Buenos Aires (IBioBA), CONICET, Partner Institute of the Max Planck Society, Buenos Aires, Argentina
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13
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Arumugam T, Adimulam T, Gokul A, Ramsuran V. Variation within the non-coding genome influences genetic and epigenetic regulation of the human leukocyte antigen genes. Front Immunol 2024; 15:1422834. [PMID: 39355248 PMCID: PMC11442197 DOI: 10.3389/fimmu.2024.1422834] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2024] [Accepted: 08/26/2024] [Indexed: 10/03/2024] Open
Abstract
Variation within the non-coding genome may influence the regulation and expression of important genes involved in immune control such as the human leukocyte antigen (HLA) system. Class I and Class II HLA molecules are essential for peptide presentation which is required for T lymphocyte activation. Single nucleotide polymorphisms within non-coding regions of HLA Class I and Class II genes may influence the expression of these genes by affecting the binding of transcription factors and chromatin modeling molecules. Furthermore, an interplay between genetic and epigenetic factors may also influence HLA expression. Epigenetic factors such as DNA methylation and non-coding RNA, regulate gene expression without changing the DNA sequence. However, genetic variation may promote or allow genes to escape regulation by epigenetic factors, resulting in altered expression. The HLA system is central to most diseases, therefore, understanding the role of genetics and epigenetics on HLA regulation will tremendously impact healthcare. The knowledge gained from these studies may lead to novel and cost-effective diagnostic approaches and therapeutic interventions. This review discusses the role of non-coding variants on HLA regulation. Furthermore, we discuss the interplay between genetic and epigenetic factors on the regulation of HLA by evaluating literature based on polymorphisms within DNA methylation and miRNA regulatory sites within class I and Class II HLA genes. We also provide insight into the importance of the HLA non-coding genome on disease, discuss ethnic-specific differences across the HLA region and provide guidelines for future HLA studies.
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Affiliation(s)
- Thilona Arumugam
- School of Laboratory Medicine, Medical Science, University of KwaZulu-Natal, Durban, South Africa
| | - Theolan Adimulam
- School of Laboratory Medicine, Medical Science, University of KwaZulu-Natal, Durban, South Africa
| | - Anmol Gokul
- School of Laboratory Medicine, Medical Science, University of KwaZulu-Natal, Durban, South Africa
| | - Veron Ramsuran
- School of Laboratory Medicine, Medical Science, University of KwaZulu-Natal, Durban, South Africa
- Centre for the AIDS Programme of Research in South Africa (CAPRISA), Durban, South Africa
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14
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Chowdhury MAR, Haq MM, Lee JH, Jeong S. Multi-faceted regulation of CREB family transcription factors. Front Mol Neurosci 2024; 17:1408949. [PMID: 39165717 PMCID: PMC11333461 DOI: 10.3389/fnmol.2024.1408949] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2024] [Accepted: 07/12/2024] [Indexed: 08/22/2024] Open
Abstract
cAMP response element-binding protein (CREB) is a ubiquitously expressed nuclear transcription factor, which can be constitutively activated regardless of external stimuli or be inducibly activated by external factors such as stressors, hormones, neurotransmitters, and growth factors. However, CREB controls diverse biological processes including cell growth, differentiation, proliferation, survival, apoptosis in a cell-type-specific manner. The diverse functions of CREB appear to be due to CREB-mediated differential gene expression that depends on cAMP response elements and multi-faceted regulation of CREB activity. Indeed, the transcriptional activity of CREB is controlled at several levels including alternative splicing, post-translational modification, dimerization, specific transcriptional co-activators, non-coding small RNAs, and epigenetic regulation. In this review, we present versatile regulatory modes of CREB family transcription factors and discuss their functional consequences.
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Affiliation(s)
- Md Arifur Rahman Chowdhury
- Department of Bioactive Material Sciences, Jeonbuk National University, Jeonju, Republic of Korea
- Department of Molecular Biology, and Research Center of Bioactive Materials, Jeonbuk National University, Jeonju, Republic of Korea
| | - Md Mazedul Haq
- Department of Bioactive Material Sciences, Jeonbuk National University, Jeonju, Republic of Korea
- Department of Molecular Biology, and Research Center of Bioactive Materials, Jeonbuk National University, Jeonju, Republic of Korea
| | - Jeong Hwan Lee
- Division of Life Sciences, Jeonbuk National University, Jeonju, Republic of Korea
| | - Sangyun Jeong
- Department of Bioactive Material Sciences, Jeonbuk National University, Jeonju, Republic of Korea
- Department of Molecular Biology, and Research Center of Bioactive Materials, Jeonbuk National University, Jeonju, Republic of Korea
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Kaimala S, Lootah SS, Mehra N, Kumar CA, Marzooqi SA, Sampath P, Ansari SA, Emerald BS. The Long Non-Coding RNA Obesity-Related (Obr) Contributes To Lipid Metabolism Through Epigenetic Regulation. ADVANCED SCIENCE (WEINHEIM, BADEN-WURTTEMBERG, GERMANY) 2024; 11:e2401939. [PMID: 38704700 PMCID: PMC11234455 DOI: 10.1002/advs.202401939] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/23/2024] [Indexed: 05/07/2024]
Abstract
Obesity is a multifactorial disease that is part of today's epidemic and also increases the risk of other metabolic diseases. Long noncoding RNAs (lncRNAs) provide one tier of regulatory mechanisms to maintain metabolic homeostasis. Although lncRNAs are a significant constituent of the mammalian genome, studies aimed at their metabolic significance, including obesity, are only beginning to be addressed. Here, a developmentally regulated lncRNA, termed as obesity related (Obr), whose expression in metabolically relevant tissues such as skeletal muscle, liver, and pancreas is altered in diet-induced obesity, is identified. The Clone 9 cell line and high-fat diet-induced obese Wistar rats are used as a model system to verify the function of Obr. By using stable expression and antisense oligonucleotide-mediated downregulation of the expression of Obr followed by different molecular biology experiments, its role in lipid metabolism is verified. It is shown that Obr associates with the cAMP response element-binding protein (Creb) and activates different transcription factors involved in lipid metabolism. Its association with the Creb histone acetyltransferase complex, which includes the cAMP response element-binding protein (CBP) and p300, positively regulates the transcription of genes involved in lipid metabolism. In addition, Obr is regulated by Pparγ in response to lipid accumulation.
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Affiliation(s)
- Suneesh Kaimala
- Department of Anatomy, College of Medicine and Health Sciences, UAE University, Al Ain, P.O. Box 15551, UAE
| | - Shareena Saeed Lootah
- Department of Anatomy, College of Medicine and Health Sciences, UAE University, Al Ain, P.O. Box 15551, UAE
| | - Neha Mehra
- Department of Anatomy, College of Medicine and Health Sciences, UAE University, Al Ain, P.O. Box 15551, UAE
| | - Challagandla Anil Kumar
- Department of Anatomy, College of Medicine and Health Sciences, UAE University, Al Ain, P.O. Box 15551, UAE
| | - Saeeda Al Marzooqi
- Department of Pathology, College of Medicine and Health Sciences, United Arab Emirates University, Al Ain, Abu Dhabi, P.O. Box 15551, UAE
| | - Prabha Sampath
- A*STAR Skin Research Laboratory, Agency for Science Technology & Research (A*STAR), Singapore, 138648, Singapore
- Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, 8 College Road, Singapore, 169857, Singapore
- Genome Institute of Singapore, Agency for Science Technology & Research (A*STAR), Singapore, 138672, Singapore
| | - Suraiya Anjum Ansari
- Department of Biochemistry and Molecular Biology, College of Medicine and Health Sciences, United Arab Emirates University, Al Ain, Abu Dhabi, P.O. Box 15551, UAE
- Zayed Center for Health Sciences, United Arab Emirates University, Al Ain, Abu Dhabi, P.O. Box 15551, UAE
- ASPIRE Precision Medicine, Research Institute Abu Dhabi, Al Ain, Abu Dhabi, P.O. Box 15551, UAE
| | - Bright Starling Emerald
- Department of Anatomy, College of Medicine and Health Sciences, UAE University, Al Ain, P.O. Box 15551, UAE
- Zayed Center for Health Sciences, United Arab Emirates University, Al Ain, Abu Dhabi, P.O. Box 15551, UAE
- ASPIRE Precision Medicine, Research Institute Abu Dhabi, Al Ain, Abu Dhabi, P.O. Box 15551, UAE
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16
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Pan AL, Audrain M, Sakakibara E, Joshi R, Zhu X, Wang Q, Wang M, Beckmann ND, Schadt EE, Gandy S, Zhang B, Ehrlich ME, Salton SR. Dual-specificity protein phosphatase 6 (DUSP6) overexpression reduces amyloid load and improves memory deficits in male 5xFAD mice. Front Aging Neurosci 2024; 16:1400447. [PMID: 39006222 PMCID: PMC11239576 DOI: 10.3389/fnagi.2024.1400447] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/13/2024] [Accepted: 06/14/2024] [Indexed: 07/16/2024] Open
Abstract
Introduction Dual specificity protein phosphatase 6 (DUSP6) was recently identified as a key hub gene in a causal VGF gene network that regulates late-onset Alzheimer's disease (AD). Importantly, decreased DUSP6 levels are correlated with an increased clinical dementia rating (CDR) in human subjects, and DUSP6 levels are additionally decreased in the 5xFAD amyloidopathy mouse model. Methods To investigate the role of DUSP6 in AD, we stereotactically injected AAV5-DUSP6 or AAV5-GFP (control) into the dorsal hippocampus (dHc) of both female and male 5xFAD or wild type mice, to induce overexpression of DUSP6 or GFP. Results Barnes maze testing indicated that DUSP6 overexpression in the dHc of 5xFAD mice improved memory deficits and was associated with reduced amyloid plaque load, Aß1-40 and Aß1-42 levels, and amyloid precursor protein processing enzyme BACE1, in male but not in female mice. Microglial activation, which was increased in 5xFAD mice, was significantly reduced by dHc DUSP6 overexpression in both males and females, as was the number of "microglial clusters," which correlated with reduced amyloid plaque size. Transcriptomic profiling of female 5xFAD hippocampus revealed upregulation of inflammatory and extracellular signal-regulated kinase pathways, while dHc DUSP6 overexpression in female 5xFAD mice downregulated a subset of genes in these pathways. Gene ontology analysis of DEGs (p < 0.05) identified a greater number of synaptic pathways that were regulated by DUSP6 overexpression in male compared to female 5xFAD. Discussion In summary, DUSP6 overexpression in dHc reduced amyloid deposition and memory deficits in male but not female 5xFAD mice, whereas reduced neuroinflammation and microglial activation were observed in both males and females, suggesting that DUSP6-induced reduction of microglial activation did not contribute to sex-dependent improvement in memory deficits. The sex-dependent regulation of synaptic pathways by DUSP6 overexpression, however, correlated with the improvement of spatial memory deficits in male but not female 5xFAD.
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Affiliation(s)
- Allen L. Pan
- Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Mickael Audrain
- Department of Neurology, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Emmy Sakakibara
- Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Rajeev Joshi
- Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Xiaodong Zhu
- Department of Psychiatry, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Qian Wang
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Mount Sinai Center for Transformative Disease Modeling, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Minghui Wang
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Mount Sinai Center for Transformative Disease Modeling, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Noam D. Beckmann
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Eric E. Schadt
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Sam Gandy
- Department of Neurology, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Department of Psychiatry and Alzheimer’s Disease Research Center, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Bin Zhang
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Mount Sinai Center for Transformative Disease Modeling, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Michelle E. Ehrlich
- Department of Neurology, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York, NY, United States
| | - Stephen R. Salton
- Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, United States
- Brookdale Department of Geriatrics and Palliative Medicine, Icahn School of Medicine at Mount Sinai, New York, NY, United States
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Zhang T, Fu W, Liu D, He Y, Wang J, Ma T. ADENOSINE INFLUENCES FOXP3 EXPRESSION OF T REGS VIA THE A2AR/CREB PATHWAY IN A MOUSE MODEL OF SEPSIS. Shock 2024; 61:924-933. [PMID: 38010286 DOI: 10.1097/shk.0000000000002281] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2023]
Abstract
ABSTRACT The adenosine concentration and forkhead box protein (Foxp3) expression in T regulatory cells (T regs ) are increased during sepsis. However, the mechanism by which adenosine induces Foxp3 expression is incompletely understood. A cecal ligation and puncture (CLP) model was constructed using C57BL/J mice. The plasma adenosine concentration and Foxp3 expression in splenic T regs were increased consistently for 15 days after sepsis onset. Analysis of the mean fluorescence intensity of Foxp3 and adenosine concentration in the same mice revealed a linear correlation. In the CLP model, adenosine 2a receptor (A2aR) blockade inhibited Foxp3 expression in T regs . In vitro activation of A2aR promoted Foxp3 expression in T regs and facilitated secretion of extracellular vesicles. Transcriptome sequencing revealed that A2aR blockade led to changes in cyclic adenosine monophosphate response element-binding protein (CREB) transcription in T regs in our sepsis model. Use of adenosine or A2aR agonists promoted CREB expression, CREB phosphorylation at S133, T reg expression of Foxp3, and enhanced inhibition of proliferation of cluster of differentiation (CD)4+ lymphocytes. A2aR blockade or inhibition of CREB expression inhibited Foxp3 expression in T regs . In the CLP model, use of CREB inhibitors could inhibit Foxp3 expression and reduce the bacterial load. In summary, adenosine in sepsis promotes CREB phosphorylation via A2aR which, in turn, upregulates Foxp3 expression in T regs .
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Affiliation(s)
- Teng Zhang
- Department of General Surgery, Tianjin Medical University General Hospital, Tianjin, China
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18
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Peng Y, Yu J, Liu F, Tang L, Li B, Zhang W, Chen K, Zhang H, Wei Y, Ma X, Shi H. Accumulation of TOX high mobility group box family member 3 promotes the oncogenesis and development of hepatocellular carcinoma through the MAPK signaling pathway. MedComm (Beijing) 2024; 5:e510. [PMID: 38463397 PMCID: PMC10924639 DOI: 10.1002/mco2.510] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2023] [Revised: 02/12/2024] [Accepted: 02/12/2024] [Indexed: 03/12/2024] Open
Abstract
Microvascular invasion (MVI) has been widely valued in the field of liver surgery because MVI positivity indicates poor prognosis in hepatocellular carcinoma (HCC) patients. However, the potential molecular mechanism underlying the poor prognosis of MVI-positive HCC patients is unclear. Therefore, this study focused on identifying the key genes leading to poor prognosis in patients with a high degree of malignancy of HCC by examining the molecular signaling pathways in MVI-positive HCC patients. Through RNA sequencing, TOX high mobility group box family member 3 (TOX3) was demonstrated to be significantly highly expressed in MVI-positive HCC tissues, which was associated with poor prognosis. The results of in vivo and in vitro showed that TOX3 can promote the oncogenesis and development of HCC by targeting key molecules of the MAPK and EMT signaling pathways. The IP-MS results indicated that proteasome degradation of TOX3 in HCC cells is potentially mediated by a tripartite motif containing 56 (TRIM56, an E3 ligase) in HCC cells. Inhibiting TRIM56 enhances TOX3 protein levels. Overall, our study identified TOX3 as a key gene in the MAPK and EMT signaling pathways in HCC, and its overexpression confers significant proliferation and invasiveness to tumor cells.
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Affiliation(s)
- Yufu Peng
- Division of Liver SurgeryDepartment of General SurgeryWest China HospitalSichuan UniversityChengduChina
- Laboratory of Integrative MedicineClinical Research Center for BreastState Key Laboratory of BiotherapyWest China HospitalSichuan University and Collaborative Innovation CenterChengduChina
| | - Jing Yu
- Laboratory of Integrative MedicineClinical Research Center for BreastState Key Laboratory of BiotherapyWest China HospitalSichuan University and Collaborative Innovation CenterChengduChina
| | - Fei Liu
- Division of Liver SurgeryDepartment of General SurgeryWest China HospitalSichuan UniversityChengduChina
| | - Leyi Tang
- Laboratory of Integrative MedicineClinical Research Center for BreastState Key Laboratory of BiotherapyWest China HospitalSichuan University and Collaborative Innovation CenterChengduChina
| | - Bo Li
- Division of Liver SurgeryDepartment of General SurgeryWest China HospitalSichuan UniversityChengduChina
| | - Wei Zhang
- Department of Critical Care MedicineState Key Laboratory of Biotherapy and Cancer CenterWest China HospitalSichuan University, China
| | - Kefei Chen
- Division of Liver SurgeryDepartment of General SurgeryWest China HospitalSichuan UniversityChengduChina
| | - Haili Zhang
- Division of Liver SurgeryDepartment of General SurgeryWest China HospitalSichuan UniversityChengduChina
| | - Yonggang Wei
- Division of Liver SurgeryDepartment of General SurgeryWest China HospitalSichuan UniversityChengduChina
| | - Xuelei Ma
- Department of BiotherapyWest China Hospital and State Key Laboratory of BiotherapySichuan UniversityChengduChina
| | - Hubing Shi
- Laboratory of Integrative MedicineClinical Research Center for BreastState Key Laboratory of BiotherapyWest China HospitalSichuan University and Collaborative Innovation CenterChengduChina
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Kazanietz MG, Cooke M. Protein kinase C signaling "in" and "to" the nucleus: Master kinases in transcriptional regulation. J Biol Chem 2024; 300:105692. [PMID: 38301892 PMCID: PMC10907189 DOI: 10.1016/j.jbc.2024.105692] [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: 10/23/2023] [Revised: 01/08/2024] [Accepted: 01/09/2024] [Indexed: 02/03/2024] Open
Abstract
PKC is a multifunctional family of Ser-Thr kinases widely implicated in the regulation of fundamental cellular functions, including proliferation, polarity, motility, and differentiation. Notwithstanding their primary cytoplasmic localization and stringent activation by cell surface receptors, PKC isozymes impel prominent nuclear signaling ultimately impacting gene expression. While transcriptional regulation may be wielded by nuclear PKCs, it most often relies on cytoplasmic phosphorylation events that result in nuclear shuttling of PKC downstream effectors, including transcription factors. As expected from the unique coupling of PKC isozymes to signaling effector pathways, glaring disparities in gene activation/repression are observed upon targeting individual PKC family members. Notably, specific PKCs control the expression and activation of transcription factors implicated in cell cycle/mitogenesis, epithelial-to-mesenchymal transition and immune function. Additionally, PKCs isozymes tightly regulate transcription factors involved in stepwise differentiation of pluripotent stem cells toward specific epithelial, mesenchymal, and hematopoietic cell lineages. Aberrant PKC expression and/or activation in pathological conditions, such as in cancer, leads to profound alterations in gene expression, leading to an extensive rewiring of transcriptional networks associated with mitogenesis, invasiveness, stemness, and tumor microenvironment dysregulation. In this review, we outline the current understanding of PKC signaling "in" and "to" the nucleus, with significant focus on established paradigms of PKC-mediated transcriptional control. Dissecting these complexities would allow the identification of relevant molecular targets implicated in a wide spectrum of diseases.
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Affiliation(s)
- Marcelo G Kazanietz
- Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
| | - Mariana Cooke
- Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
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Dinevska M, Widodo SS, Cook L, Stylli SS, Ramsay RG, Mantamadiotis T. CREB: A multifaceted transcriptional regulator of neural and immune function in CNS tumors. Brain Behav Immun 2024; 116:140-149. [PMID: 38070619 DOI: 10.1016/j.bbi.2023.12.002] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/06/2023] [Revised: 11/16/2023] [Accepted: 12/04/2023] [Indexed: 01/21/2024] Open
Abstract
Cancers of the central nervous system (CNS) are unique with respect to their tumor microenvironment. Such a status is due to immune-privilege and the cellular behaviors within a highly networked, neural-rich milieu. During tumor development in the CNS, neural, immune and cancer cells establish complex cell-to-cell communication networks which mimic physiological functions, including paracrine signaling and synapse-like formations. This crosstalk regulates diverse pathological functions contributing to tumor progression. In the CNS, regulation of physiological and pathological functions relies on various cell signaling and transcription programs. At the core of these events lies the cyclic adenosine monophosphate (cAMP) response element binding protein (CREB), a master transcriptional regulator in the CNS. CREB is a kinase inducible transcription factor which regulates many CNS functions, including neurogenesis, neuronal survival, neuronal activation and long-term memory. Here, we discuss how CREB-regulated mechanisms operating in diverse cell types, which control development and function of the CNS, are co-opted in CNS tumors.
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Affiliation(s)
- Marija Dinevska
- Department of Surgery, Royal Melbourne Hospital, The University of Melbourne, Parkville, VIC, Australia
| | - Samuel S Widodo
- Department of Surgery, Royal Melbourne Hospital, The University of Melbourne, Parkville, VIC, Australia
| | - Laura Cook
- Department of Microbiology and Immunology, The University of Melbourne at The Peter Doherty Institute for Infection and Immunity, Parkville, VIC, Australia
| | - Stanley S Stylli
- Department of Surgery, Royal Melbourne Hospital, The University of Melbourne, Parkville, VIC, Australia; Department of Neurosurgery, Royal Melbourne Hospital, Parkville, VIC, Australia
| | - Robert G Ramsay
- Cancer Immunology Program, Peter MacCallum Cancer Centre, Melbourne, Australia; Sir Peter MacCallum Department of Oncology and the Department of Clinical Pathology, The University of Melbourne, Melbourne, Australia
| | - Theo Mantamadiotis
- Department of Surgery, Royal Melbourne Hospital, The University of Melbourne, Parkville, VIC, Australia; Department of Microbiology and Immunology, The University of Melbourne at The Peter Doherty Institute for Infection and Immunity, Parkville, VIC, Australia; Centre for Stem Cell Systems, The University of Melbourne, Parkville, VIC, Australia.
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21
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Pizzoni A, Zhang X, Altschuler DL. From membrane to nucleus: A three-wave hypothesis of cAMP signaling. J Biol Chem 2024; 300:105497. [PMID: 38016514 PMCID: PMC10788541 DOI: 10.1016/j.jbc.2023.105497] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/18/2023] [Revised: 11/14/2023] [Accepted: 11/19/2023] [Indexed: 11/30/2023] Open
Abstract
For many decades, our understanding of G protein-coupled receptor (GPCR) activity and cyclic AMP (cAMP) signaling was limited exclusively to the plasma membrane. However, a growing body of evidence has challenged this view by introducing the concept of endocytosis-dependent GPCR signaling. This emerging paradigm emphasizes not only the sustained production of cAMP but also its precise subcellular localization, thus transforming our understanding of the spatiotemporal organization of this process. Starting from this alternative point of view, our recent work sheds light on the role of an endocytosis-dependent calcium release from the endoplasmic reticulum in the control of nuclear cAMP levels. This is achieved through the activation of local soluble adenylyl cyclase, which in turn regulates the activation of local protein kinase A (PKA) and downstream transcriptional events. In this review, we explore the dynamic evolution of research on cyclic AMP signaling, including the findings that led us to formulate the novel three-wave hypothesis. We delve into how we abandoned the paradigm of cAMP generation limited to the plasma membrane and the changing perspectives on the rate-limiting step in nuclear PKA activation.
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Affiliation(s)
- Alejandro Pizzoni
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Xuefeng Zhang
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
| | - Daniel L Altschuler
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
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Ozenberger BB, Li L, Wilson ER, Lazar AJ, Barrott JJ, Jones KB. EWSR1::ATF1 Orchestrates the Clear Cell Sarcoma Transcriptome in Human Tumors and a Mouse Genetic Model. Cancers (Basel) 2023; 15:5750. [PMID: 38136296 PMCID: PMC10742207 DOI: 10.3390/cancers15245750] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/01/2023] [Revised: 12/03/2023] [Accepted: 12/04/2023] [Indexed: 12/24/2023] Open
Abstract
Clear cell sarcoma (CCS) is a rare, aggressive malignancy that most frequently arises in the soft tissues of the extremities. It is defined and driven by expression of one member of a family of related translocation-generated fusion oncogenes, the most common of which is EWSR1::ATF1. The EWSR1::ATF1 fusion oncoprotein reprograms transcription. However, the binding distribution of EWSR1::ATF1 across the genome and its target genes remain unclear. Here, we interrogated the genomic distribution of V5-tagged EWSR1::ATF1 in tumors it had induced upon expression in mice that also recapitulated the transcriptome of human CCS. ChIP-sequencing of V5-EWSR1::ATF1 identified previously unreported motifs including the AP1 motif and motif comprised of TGA repeats that resemble GGAA-repeating microsatellites bound by EWSR1::FLI1 in Ewing sarcoma. ChIP-sequencing of H3K27ac identified super enhancers in the mouse model and human contexts of CCS, which showed a shared super enhancer structure that associates with activated genes.
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Affiliation(s)
- Benjamin B. Ozenberger
- Department of Oncological Sciences, University of Utah School of Medicine, Salt Lake City, UT 84132, USA; (B.B.O.); (L.L.); (E.R.W.)
- Department of Orthopaedics, University of Utah School of Medicine, Salt Lake City, UT 84132, USA
| | - Li Li
- Department of Oncological Sciences, University of Utah School of Medicine, Salt Lake City, UT 84132, USA; (B.B.O.); (L.L.); (E.R.W.)
- Department of Orthopaedics, University of Utah School of Medicine, Salt Lake City, UT 84132, USA
| | - Emily R. Wilson
- Department of Oncological Sciences, University of Utah School of Medicine, Salt Lake City, UT 84132, USA; (B.B.O.); (L.L.); (E.R.W.)
| | - Alexander J. Lazar
- Department of Pathology, Genomic Medicine and Dermatology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA;
| | - Jared J. Barrott
- Department of Biology, Brigham Young University, Provo, UT 84602, USA;
| | - Kevin B. Jones
- Department of Oncological Sciences, University of Utah School of Medicine, Salt Lake City, UT 84132, USA; (B.B.O.); (L.L.); (E.R.W.)
- Department of Orthopaedics, University of Utah School of Medicine, Salt Lake City, UT 84132, USA
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23
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Shan G, Bi G, Zhao G, Liang J, Bian Y, Zhang H, Jin X, Hu Z, Yao G, Fan H, Zhan C. Inhibition of PKA/CREB1 pathway confers sensitivity to ferroptosis in non-small cell lung cancer. Respir Res 2023; 24:277. [PMID: 37957645 PMCID: PMC10644539 DOI: 10.1186/s12931-023-02567-3] [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: 07/21/2023] [Accepted: 10/17/2023] [Indexed: 11/15/2023] Open
Abstract
Ferroptosis is a type of regulated cell death characterized by iron accumulation and lipid peroxidation. The molecular mechanisms underlying ferroptosis regulation in non-small cell lung cancer (NSCLC) are poorly understood. In this study, we found that protein kinase A (PKA) inhibition enhanced ferroptosis susceptibility in NSCLC cells, as evidenced by reduced cell viability and increased lipid peroxidation. We further identified cAMP-responsive element protein 1 (CREB1), a transcription factor and a substrate of PKA, as a key regulator of ferroptosis. Knockdown of CREB1 sensitized NSCLC cells to ferroptosis inducers (FINs) and abolished the effects of PKA inhibitor and agonist, revealing the pivotal role of CREB1 in ferroptosis regulation. Using a high-throughput screening approach and subsequent validation by chromatin immunoprecipitation (ChIP) and dual-luciferase assays, we discovered that CREB1 transcriptionally activated stearoyl-CoA desaturase (SCD), an enzyme that catalyzes the conversion of saturated fatty acids to monounsaturated fatty acids. SCD conferred ferroptosis resistance by decreasing the availability of polyunsaturated fatty acids for lipid peroxidation, and its overexpression rescued the effect of CREB1 knockdown on ferroptosis in vitro. Besides, CREB1 knockdown suppressed xenograft tumor growth in the presence of Imidazole Ketone Erastin (IKE), a potent FIN, and this effect was reversed by SCD. Finally, we showed that high expression of CREB1 was associated with poor prognosis in NSCLC patients from public datasets and our institution. Collectively, this study illustrates the effect of PKA/CREB1/SCD axis in regulating ferroptosis of NSCLC, targeting this pathway may provide new strategies for treating NSCLC patients.
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Affiliation(s)
- Guangyao Shan
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Guoshu Bi
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Guangyin Zhao
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Jiaqi Liang
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Yunyi Bian
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Huan Zhang
- Division of Thoracic Surgery, School of Medicine, Sichuan Cancer Hospital and Research Institute, University of Electronic Science and Technology of China (UESTC), Chengdu, China
| | - Xing Jin
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Zhengyang Hu
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Guangyu Yao
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China
| | - Hong Fan
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China.
| | - Cheng Zhan
- Department of Thoracic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China.
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Nie J, Ling Y, Jin M, Chen Z, Liu W, Shen W, Fang T, Li J, He Y. Butyrate enhances erastin-induced ferroptosis of osteosarcoma cells via regulating ATF3/SLC7A11 pathway. Eur J Pharmacol 2023; 957:176009. [PMID: 37619784 DOI: 10.1016/j.ejphar.2023.176009] [Citation(s) in RCA: 12] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2022] [Revised: 08/04/2023] [Accepted: 08/21/2023] [Indexed: 08/26/2023]
Abstract
Osteosarcoma (OS) is a highly fatal bone tumor characterized by high degree of malignancy and early lung metastasis. Traditional chemotherapy fails in improving the efficacy and survival rate of patients with OS. Butyrate (NaBu) has been reported as a new antitumor drug for inhibiting proliferation and inducing apoptosis in various cancer cells. However, the effect of NaBu on the ferroptosis of OS is still unknown. This study aimed to investigate whether NaBu promotes erastin-induced ferroptosis in OS cells and to uncover the underlying mechanism. Here, we found that NaBu significantly enhanced erastin-induced ferroptosis in vitro and in vivo. Compared with the group that erastin used alonely, pre-treating with NaBu exacerbated erastin-meditated GSH depletion, lipid peroxidation, and mitochondrial morphologic changes in OS cells. In a subcutaneous OS model, NaBu combined with erastin significantly reduced tumor growth and increased the levels of 4-HNE. Mechanistically, NaBu downregulated SLC7A11 transcription via regulating ATF3 expression. Overexpression of ATF3 facilitated erastin to induce ferroptosis, while ATF3 knockdown attenuated NaBu-induced ferroptosis sensitivity. In conclusion, our findings revealed a previously unidentified role of NaBu in erastin-induced ferroptosis by regulating SLC7A11, suggesting that NaBu may be a potential therapeutic agent for OS treatment.
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Affiliation(s)
- Jiangbo Nie
- Department of Orthopedics, Huzhou Central Hospital, Huzhou, Zhejiang, 313000, China; Zhejiang University Huzhou Hospital, Huzhou, Zhejiang, 313000, China
| | - Yuhang Ling
- Central Laboratory, The First Affiliated Hospital of Huzhou University, Huzhou, Zhejiang, 313000, China; Huzhou Key Laboratory of Translational Medicine, The First Affiliated Hospital of Huzhou University, Huzhou, Zhejiang, 313000, China
| | - Mingchao Jin
- Department of Orthopedics, Huzhou Central Hospital, Huzhou, Zhejiang, 313000, China; Zhejiang University Huzhou Hospital, Huzhou, Zhejiang, 313000, China
| | - Zhuo Chen
- Department of Orthopedics, Huzhou Central Hospital, Huzhou, Zhejiang, 313000, China; Zhejiang University Huzhou Hospital, Huzhou, Zhejiang, 313000, China
| | - Wei Liu
- Department of Orthopedics, Huzhou Central Hospital, Huzhou, Zhejiang, 313000, China
| | - Weiyun Shen
- Central Laboratory, The First Affiliated Hospital of Huzhou University, Huzhou, Zhejiang, 313000, China; Huzhou Key Laboratory of Translational Medicine, The First Affiliated Hospital of Huzhou University, Huzhou, Zhejiang, 313000, China
| | - Tianshun Fang
- Department of Orthopedics, Huzhou Central Hospital, Huzhou, Zhejiang, 313000, China; Zhejiang University Huzhou Hospital, Huzhou, Zhejiang, 313000, China
| | - Jianyou Li
- Department of Orthopedics, Huzhou Central Hospital, Huzhou, Zhejiang, 313000, China; Zhejiang University Huzhou Hospital, Huzhou, Zhejiang, 313000, China.
| | - Ying He
- Central Laboratory, The First Affiliated Hospital of Huzhou University, Huzhou, Zhejiang, 313000, China; Huzhou Key Laboratory of Translational Medicine, The First Affiliated Hospital of Huzhou University, Huzhou, Zhejiang, 313000, China.
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25
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Lu X, Zhong L, Lindell E, Veanes M, Guo J, Zhao M, Salehi M, Swartling FJ, Chen X, Sjöblom T, Zhang X. Identification of ATF3 as a novel protective signature of quiescent colorectal tumor cells. Cell Death Dis 2023; 14:676. [PMID: 37833290 PMCID: PMC10576032 DOI: 10.1038/s41419-023-06204-1] [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: 07/06/2023] [Revised: 09/20/2023] [Accepted: 09/28/2023] [Indexed: 10/15/2023]
Abstract
Colorectal cancer (CRC) is the third most common cancer and the second leading cause of death in the world. In most cases, drug resistance and tumor recurrence are ultimately inevitable. One obstacle is the presence of chemotherapy-insensitive quiescent cancer cells (QCCs). Identification of unique features of QCCs may facilitate the development of new targeted therapeutic strategies to eliminate tumor cells and thereby delay tumor recurrence. Here, using single-cell RNA sequencing, we classified proliferating and quiescent cancer cell populations in the human colorectal cancer spheroid model and identified ATF3 as a novel signature of QCCs that could support cells living in a metabolically restricted microenvironment. RNA velocity further showed a shift from the QCC group to the PCC group indicating the regenerative capacity of the QCCs. Our further results of epigenetic analysis, STING analysis, and evaluation of TCGA COAD datasets build a conclusion that ATF3 can interact with DDIT4 and TRIB3 at the transcriptional level. In addition, decreasing the expression level of ATF3 could enhance the efficacy of 5-FU on CRC MCTS models. In conclusion, ATF3 was identified as a novel marker of QCCs, and combining conventional drugs targeting PCCs with an option to target QCCs by reducing ATF3 expression levels may be a promising strategy for more efficient removal of tumor cells.
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Affiliation(s)
- Xi Lu
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Lei Zhong
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
- Department of Pharmacy, Personalized Drug Therapy Key Laboratory of Sichuan Province, Sichuan Provincial People's Hospital, Sichuan, China
| | - Emma Lindell
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Margus Veanes
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Jing Guo
- Centre for Computational Biology, Duke-NUS Medical School, 8 College Road, 169857, Singapore, Singapore
| | - Miao Zhao
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Maede Salehi
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Fredrik J Swartling
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Xingqi Chen
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Tobias Sjöblom
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden
| | - Xiaonan Zhang
- Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden.
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26
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Meurer F, Häberlein H, Franken S. Ivy Leaf Dry Extract EA 575 ® Has an Inhibitory Effect on the Signalling Cascade of Adenosine Receptor A 2B. Int J Mol Sci 2023; 24:12373. [PMID: 37569749 PMCID: PMC10418604 DOI: 10.3390/ijms241512373] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/07/2023] [Revised: 07/26/2023] [Accepted: 07/26/2023] [Indexed: 08/13/2023] Open
Abstract
Ivy leaf dry extract EA 575® is used to improve complaints of chronic inflammatory bronchial diseases and acute inflammation of the respiratory tract accompanied by coughing. Its mechanism of action has so far been explained by influencing β2-adrenergic signal transduction. In the present study, we investigated a possible influence on adenosine receptor A2B (A2BAR) signalling, as it has been described to play a significant and detrimental role in chronic inflammatory airway diseases. The influence of EA 575® on A2BAR signalling was assessed with measurements of dynamic mass redistribution. Subsequently, the effects on A2BAR-mediated second messenger cAMP levels, β-arrestin 2 recruitment, and cAMP response element (CRE) activation were examined using luciferase-based HEK293 reporter cell lines. Lastly, the impact on A2BAR-mediated IL-6 release in Calu-3 epithelial lung cells was investigated via the Lumit™ Immunoassay. Additionally, the adenosine receptor subtype mediating these effects was specified, and A2BAR was found to be responsible. The present study demonstrates an inhibitory influence of EA 575® on A2BAR-mediated general cellular response, cAMP levels, β-arrestin 2 recruitment, CRE activation, and IL-6 release. Since these EA 575®-mediated effects occur within a time frame of several hours of incubation, its mode of action can be described as indirect. The present data are the first to describe an inhibitory effect of EA 575® on A2BAR signalling. This may offer an explanation for the beneficial clinical effects of the extract in adjuvant asthma therapy.
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Affiliation(s)
| | | | - Sebastian Franken
- Institute of Biochemistry and Molecular Biology, Medical Faculty, University of Bonn, 53115 Bonn, Germany; (F.M.); (H.H.)
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27
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Li Y, Patterson MR, Morgan EL, Wasson CW, Ryder EL, Barba‐Moreno D, Scarth JA, Wang M, Macdonald A. CREB1 activation promotes human papillomavirus oncogene expression and cervical cancer cell transformation. J Med Virol 2023; 95:e29025. [PMID: 37565725 PMCID: PMC10952218 DOI: 10.1002/jmv.29025] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2023] [Revised: 07/03/2023] [Accepted: 07/10/2023] [Indexed: 08/12/2023]
Abstract
Human papillomaviruses (HPVs) infect the oral and anogenital mucosa and can cause cancer. The high-risk (HR)-HPV oncoproteins, E6 and E7, hijack cellular factors to promote cell proliferation, delay differentiation and induce genomic instability, thus predisposing infected cells to malignant transformation. cAMP response element (CRE)-binding protein 1 (CREB1) is a master transcription factor that can function as a proto-oncogene, the abnormal activity of which is associated with multiple cancers. However, little is known about the interplay between HPV and CREB1 activity in cervical cancer or the productive HPV lifecycle. We show that CREB is activated in productively infected primary keratinocytes and that CREB1 expression and phosphorylation is associated with the progression of HPV+ cervical disease. The depletion of CREB1 or inhibition of CREB1 activity results in decreased cell proliferation and reduced expression of markers of epithelial to mesenchymal transition, coupled with reduced migration in HPV+ cervical cancer cell lines. CREB1 expression is negatively regulated by the tumor suppressor microRNA, miR-203a, and CREB1 phosphorylation is controlled through the MAPK/MSK pathway. Crucially, CREB1 directly binds the viral promoter to upregulate transcription of the E6/E7 oncogenes, establishing a positive feedback loop between the HPV oncoproteins and CREB1. Our findings demonstrate the oncogenic function of CREB1 in HPV+ cervical cancer and its relationship with the HPV oncogenes.
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Affiliation(s)
- Yigen Li
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
| | - Molly R. Patterson
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
| | | | - Christopher W. Wasson
- Leeds Institute of Rheumatic and Musculoskeletal Medicine, Faculty of Medicine and HealthUniversity of LeedsLeedsWest YorkshireUK
| | - Emma L. Ryder
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
| | - Diego Barba‐Moreno
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
| | - James A. Scarth
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
| | - Miao Wang
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
| | - Andrew Macdonald
- School of Molecular and Cellular Biology, Faculty of Biological SciencesUniversity of LeedsLeedsWest YorkshireUK
- Astbury Centre for Structural Molecular BiologyUniversity of LeedsLeedsWest YorkshireUK
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Mukherjee S, Sarkar AK, Lahiri A, Sengupta Bandyopadhyay S. Analysis of the interaction of a non-canonical twin half-site of Cyclic AMP-Response Element (CRE) with CRE-binding protein. Biochimie 2023; 211:25-34. [PMID: 36842626 DOI: 10.1016/j.biochi.2023.02.011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2022] [Revised: 12/23/2022] [Accepted: 02/17/2023] [Indexed: 02/26/2023]
Abstract
Differential regulation of a gene having either canonical or non-canonical cyclic AMP response element (CRE) in its promoter is primarily accomplished by its interactions with CREB (cAMP-response element binding protein). The present study aims to delineate the mechanism of the CREB-CRE interactions at the Oncostatin-M (osm) promoter by in vitro and in silico approaches. The non-canonical CREosm consists of two half-CREs separated by a short intervening sequence of 9 base pairs. In this study, in vitro binding assays revealed that out of the two CRE half-sites, the right half-CRE was indispensable for binding of CREB, while the left sequence showed weaker binding ability and specificity. Genome-wide modeling and high throughput free energy calculations for the energy-minimized models containing CREB-CREosm revealed that there was no difference in the binding of CREB to the right half of CREosm site when compared to the entire CREosm. These results were in accordance with the in vitro studies, confirming the indispensable role of the right half-CREosm site in stable complex formation with the CREB protein. Additionally, conversion of the right half-CREosm site to a canonical CRE palindrome showed stronger CREB binding, irrespective of the presence or absence of the left CRE sequence. Thus, the present study establishes an interesting insight into the interaction of CREB with a CRE variant located at the far end of a TATA-less promoter of a cytokine-encoding gene, which in turn could be involved in the regulation of transcription under specific conditions.
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Affiliation(s)
- Srimoyee Mukherjee
- Department of Biophysics, Molecular Biology and Bioinformatics, University of Calcutta, 92 A.P.C. Road, Kolkata, 700009, India
| | - Aditya Kumar Sarkar
- Department of Biophysics, Molecular Biology and Bioinformatics, University of Calcutta, 92 A.P.C. Road, Kolkata, 700009, India
| | - Ansuman Lahiri
- Department of Biophysics, Molecular Biology and Bioinformatics, University of Calcutta, 92 A.P.C. Road, Kolkata, 700009, India
| | - Sumita Sengupta Bandyopadhyay
- Department of Biophysics, Molecular Biology and Bioinformatics, University of Calcutta, 92 A.P.C. Road, Kolkata, 700009, India.
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29
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Tran TT, Huang WJ, Lin H, Chen HH. New Synthesized Activating Transcription Factor 3 Inducer SW20.1 Suppresses Resistin-Induced Metabolic Syndrome. Biomedicines 2023; 11:1509. [PMID: 37371606 DOI: 10.3390/biomedicines11061509] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2023] [Revised: 05/15/2023] [Accepted: 05/18/2023] [Indexed: 06/29/2023] Open
Abstract
Obesity is an emerging concern globally with increasing prevalence. Obesity is associated with many diseases, such as cardiovascular disease, dyslipidemia, and cancer. Thus, effective new antiobesity drugs should be urgently developed. We synthesized SW20.1, a compound that induces activating transcription factor 3 (ATF3) expression. The results of Oil Red O staining and quantitative real-time polymerase chain reaction revealed that SW20.1 was more effective in reducing lipid accumulation in 3T3-L1 preadipocytes than the previously synthesized ST32db, and that it inhibited the expression of the genes involved in adipogenesis and lipogenesis. A chromatin immunoprecipitation assay indicated that SW20.1 inhibited adipogenesis and lipogenesis by binding to the upstream promoter region of resistin at two sites (-2861/-2854 and -241/-234). In mice, the intraperitoneal administration of SW20.1 reduced body weight, white adipocyte weight in different regions, serum cholesterol levels, adipogenesis-related gene expression, hepatic steatosis, and serum resistin levels. Overall, SW20.1 exerts antiobesity effects by inhibiting resistin through the ATF3 pathway. Our study results indicate that SW20.1 is a promising therapeutic drug for diet-induced obesity.
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Affiliation(s)
- Tu T Tran
- International Ph.D. Program in Medicine, College of Medicine, Taipei Medical University, Taipei 110, Taiwan
- Department of Internal Medicine, Thai Nguyen University of Medicine and Pharmacy, Thai Nguyen 241-17, Vietnam
| | - Wei-Jan Huang
- School of Pharmacy, Taipei Medical University, Taipei 110, Taiwan
| | - Heng Lin
- Department of Physiology, School of Medicine, College of Medicine, Taipei Medical University, Taipei 110, Taiwan
| | - Hsi-Hsien Chen
- TMU Research Center of Urology and Kidney, Taipei Medical University, Taipei 110, Taiwan
- Division of Nephrology, Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei 110, Taiwan
- Division of Nephrology, Department of Internal Medicine, Taipei Medical University Hospital, Taipei 110, Taiwan
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Bentley EP, Scholl D, Wright PE, Deniz AA. Coupling of binding and differential subdomain folding of the intrinsically disordered transcription factor CREB. FEBS Lett 2023; 597:917-932. [PMID: 36480418 PMCID: PMC10089947 DOI: 10.1002/1873-3468.14554] [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: 08/18/2022] [Revised: 11/07/2022] [Accepted: 11/28/2022] [Indexed: 12/13/2022]
Abstract
The cyclic AMP response element binding protein (CREB) contains a basic leucine zipper motif (bZIP) that forms a coiled coil structure upon dimerization and specific DNA binding. Although this state is well characterized, key features of CREB bZIP binding and folding are not well understood. We used single-molecule Förster resonance energy transfer (smFRET) to probe conformations of CREB bZIP subdomains. We found differential folding of the basic region and leucine zipper in response to different binding partners; a strong and previously unreported DNA-independent dimerization affinity; folding upon binding to nonspecific DNA; and evidence of long-range interdomain interactions in full-length CREB that modulate DNA binding. These studies provide new insights into DNA binding and dimerization and have implications for CREB function.
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Affiliation(s)
- Emily P. Bentley
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037
| | - Daniel Scholl
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037
| | - Peter E. Wright
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037
| | - Ashok A. Deniz
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037
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Nieto-Felipe J, Macias-Diaz A, Sanchez-Collado J, Berna-Erro A, Jardin I, Salido GM, Lopez JJ, Rosado JA. Role of Orai-family channels in the activation and regulation of transcriptional activity. J Cell Physiol 2023; 238:714-726. [PMID: 36952615 DOI: 10.1002/jcp.30971] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2022] [Revised: 01/12/2023] [Accepted: 01/27/2023] [Indexed: 03/25/2023]
Abstract
Store operated Ca2+ entry (SOCE) is a cornerstone for the maintenance of intracellular Ca2+ homeostasis and the regulation of a variety of cellular functions. SOCE is mediated by STIM and Orai proteins following the activation of inositol 1,4,5-trisphosphate receptors. Then, a reduction of the endoplasmic reticulum intraluminal Ca2+ concentration is sensed by STIM proteins, which undergo a conformational change and activate plasma membrane Ca2+ channels comprised by Orai proteins. STIM1/Orai-mediated Ca2+ signals are finely regulated and modulate the activity of different transcription factors, including certain isoforms of the nuclear factor of activated T-cells, the cAMP-response element binding protein, the nuclear factor κ-light chain-enhancer of activated B cells, c-fos, and c-myc. These transcription factors associate SOCE with a plethora of signaling events and cellular functions. Here we provide an overview of the current knowledge about the role of Orai channels in the regulation of transcription factors through Ca2+ -dependent signaling pathways.
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Affiliation(s)
- Joel Nieto-Felipe
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Alvaro Macias-Diaz
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Jose Sanchez-Collado
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Alejandro Berna-Erro
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Isaac Jardin
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Gines M Salido
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Jose J Lopez
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
| | - Juan A Rosado
- Departamento de Fisiología, Instituto Universitario de Biomarcadores de Patologías Moleculares, Universidad de Extremadura, Caceres, Spain
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Qu HQ, Kao C, Garifallou J, Wang F, Snyder J, Slater DJ, Hou C, March M, Connolly JJ, Glessner JT, Hakonarson H. Single Cell Transcriptome Analysis of Peripheral Blood Mononuclear Cells in Freshly Isolated versus Stored Blood Samples. Genes (Basel) 2023; 14:142. [PMID: 36672883 PMCID: PMC9859202 DOI: 10.3390/genes14010142] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2022] [Revised: 12/27/2022] [Accepted: 01/03/2023] [Indexed: 01/06/2023] Open
Abstract
BACKGROUND Peripheral blood mononuclear cells (PBMCs) are widely used as a model in the study of different human diseases. There is often a time delay from blood collection to PBMC isolation during the sampling process, which can result in an experimental bias, particularly when performing single cell RNA-seq (scRNAseq) studies. METHODS This study examined the impact of different time periods from blood draw to PBMC isolation on the subsequent transcriptome profiling of different cell types in PBMCs by scRNAseq using the 10X Chromium Single Cell Gene Expression assay. RESULTS Examining the five major cell types constituting the PBMC cell population, i.e., CD4+ T cells, CD8+ T cells, NK cells, monocytes, and B cells, both common changes and cell-type-specific changes were observed in the single cell transcriptome profiling over time. In particular, the upregulation of genes regulated by NF-kB in response to TNF was observed in all five cell types. Significant changes in key genes involved in AP-1 signaling were also observed. RBC contamination was a major issue in stored blood, whereas RBC adherence had no direct impact on the cell transcriptome. CONCLUSIONS Significant transcriptome changes were observed across different PBMC cell types as a factor of time from blood draw to PBMC isolation and as a consequence of blood storage. This should be kept in mind when interpreting experimental results.
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Affiliation(s)
- Hui-Qi Qu
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Charlly Kao
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - James Garifallou
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Fengxiang Wang
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - James Snyder
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Diana J. Slater
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Cuiping Hou
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Michael March
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - John J. Connolly
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Joseph T. Glessner
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
- Department of Pediatrics, The Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA
- Division of Human Genetics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
| | - Hakon Hakonarson
- The Center for Applied Genomics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
- Department of Pediatrics, The Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA
- Division of Human Genetics, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
- Division of Pulmonary Medicine, Children’s Hospital of Philadelphia, Philadelphia, PA 19104, USA
- Faculty of Medicine, University of Iceland, 101 Reykjavik, Iceland
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Dexmedetomidine Attenuates Methotrexate-Induced Neurotoxicity and Memory Deficits in Rats through Improving Hippocampal Neurogenesis: The Role of miR-15a/ROCK-1/ERK1/2/CREB/BDNF Pathway Modulation. Int J Mol Sci 2023; 24:ijms24010766. [PMID: 36614208 PMCID: PMC9821704 DOI: 10.3390/ijms24010766] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2022] [Revised: 12/20/2022] [Accepted: 12/21/2022] [Indexed: 01/04/2023] Open
Abstract
Methotrexate (MTX) is a widely used neurotoxic drug with broad antineoplastic and immunosuppressant spectra. However, the exact molecular mechanisms by which MTX inhibits hippocampal neurogenesis are yet unclear. Dexmedetomidine (Dex), an α2-adrenergic receptor agonist, has recently shown neuroprotective effects; however, its full mechanism is unexplored. This study investigated the potential of Dex to mitigate MTX-induced neurotoxicity and memory impairment in rats and the possible role of the miR-15a/ROCK-1/ERK1/2/CREB/BDNF pathway. Notably, no former studies have linked this pathway to MTX-induced neurotoxicity. Male Sprague Dawley rats were placed into four groups. Group 1 received saline i.p. daily and i.v. on days 8 and 15. Group 2 received Dex at 10 μg/kg/day i.p. for 30 days. Group 3 received MTX at 75 mg/kg i.v. on days 8 and 15, followed by four i.p. doses of leucovorin at 6 mg/kg after 18 h and 3 mg/kg after 26, 42, and 50 h. Group 4 received MTX and leucovorin as in group 3 and Dex daily dosages as in group 2. Bioinformatic analysis identified the association of miR-15a with ROCK-1/ERK1/2/CREB/BDNF and neurogenesis. MTX lowered hippocampal doublecortin and Ki-67, two markers of neurogenesis. This was associated with the downregulation of miR-15a, upregulation of its target ROCK-1, and reduction in the downstream ERK1/2/CREB/BDNF pathway, along with disturbed hippocampal redox state. Novel object recognition and Morris water maze tests demonstrated the MTX-induced memory deficiencies. Dex co-treatment reversed the MTX-induced behavioral, biochemical, and histological alterations in the rats. These neuroprotective actions could be partly mediated through modulating the miR-15a/ROCK-1/ERK1/2/CREB/BDNF pathway, which enhances hippocampal neurogenesis.
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Kim D, Lee S, Cho YH, Kang MJ, Ku CR, Chi H, Ahn J, Lee K, Han J, Chi S, Song MY, Cha SH, Lee EJ. Long-acting recombinant human follicle-stimulating hormone (SAFA-FSH) enhances spermatogenesis. Front Endocrinol (Lausanne) 2023; 14:1132172. [PMID: 36909328 PMCID: PMC9996080 DOI: 10.3389/fendo.2023.1132172] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/26/2022] [Accepted: 02/13/2023] [Indexed: 02/25/2023] Open
Abstract
INTRODUCTION Administration of follicle-stimulating hormone (FSH) has been recommended to stimulate spermatogenesis in infertile men with hypogonadotropic hypogonadism, whose sperm counts do not respond to human chorionic gonadotropin alone. However, FSH has a short serum half-life requiring frequent administration to maintain its therapeutic efficacy. To improve its pharmacokinetic properties, we developed a unique albumin-binder technology, termed "anti-serum albumin Fab-associated" (SAFA) technology. We tested the feasibility of applying SAFA technology to create long-acting FSH as a therapeutic candidate for patients with hypogonadotropic hypogonadism. METHODS SAFA-FSH was produced using a Chinese hamster ovary expression system. To confirm the biological function, the production of cyclic AMP and phosphorylation of ERK and CREB were measured in TM4-FSHR cells. The effect of gonadotropin-releasing hormone agonists on spermatogenesis in a hypogonadal rat model was investigated. RESULTS In in vitro experiments, SAFA-FSH treatment increased the production of cyclic AMP and increased the phosphorylation of ERK and CREB in a dose-dependent manner. In animal experiments, sperm production was not restored by human chorionic gonadotropin treatment alone, but was restored after additional recombinant FSH treatment thrice per week or once every 5 days. Sperm production was restored even after additional SAFA-FSH treatment at intervals of once every 5 or 10 days. DISCUSSION Long-acting FSH with bioactivity was successfully created using SAFA technology. These data support further development of SAFA-FSH in a clinical setting, potentially representing an important advancement in the treatment of patients with hypogonadotropic hypogonadism.
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Affiliation(s)
- Daham Kim
- Department of Internal Medicine, Institute of Endocrine Research, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Soohyun Lee
- Department of Internal Medicine, Institute of Endocrine Research, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Yoon Hee Cho
- Department of Internal Medicine, Institute of Endocrine Research, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Min Jeong Kang
- Department of Internal Medicine, Institute of Endocrine Research, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Cheol Ryong Ku
- Department of Internal Medicine, Institute of Endocrine Research, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Hyunjin Chi
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Jungsuk Ahn
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Kyungsun Lee
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Jaekyu Han
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Susan Chi
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Moo Young Song
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Sang-Hoon Cha
- AprilBio Co., Ltd., Rm 602, Biomedical Science Building, Kangwon National University, Chuncheon, Republic of Korea
| | - Eun Jig Lee
- Department of Internal Medicine, Institute of Endocrine Research, Yonsei University College of Medicine, Seoul, Republic of Korea
- *Correspondence: Eun Jig Lee,
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Kilanowska A, Ziółkowska A, Stasiak P, Gibas-Dorna M. cAMP-Dependent Signaling and Ovarian Cancer. Cells 2022; 11:cells11233835. [PMID: 36497095 PMCID: PMC9738761 DOI: 10.3390/cells11233835] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2022] [Revised: 11/23/2022] [Accepted: 11/24/2022] [Indexed: 12/02/2022] Open
Abstract
cAMP-dependent pathway is one of the most significant signaling cascades in healthy and neoplastic ovarian cells. Working through its major effector proteins-PKA and EPAC-it regulates gene expression and many cellular functions. PKA promotes the phosphorylation of cAMP response element-binding protein (CREB) which mediates gene transcription, cell migration, mitochondrial homeostasis, cell proliferation, and death. EPAC, on the other hand, is involved in cell adhesion, binding, differentiation, and interaction between cell junctions. Ovarian cancer growth and metabolism largely depend on changes in the signal processing of the cAMP-PKA-CREB axis, often associated with neoplastic transformation, metastasis, proliferation, and inhibition of apoptosis. In addition, the intracellular level of cAMP also determines the course of other pathways including AKT, ERK, MAPK, and mTOR, that are hypo- or hyperactivated among patients with ovarian neoplasm. With this review, we summarize the current findings on cAMP signaling in the ovary and its association with carcinogenesis, multiplication, metastasis, and survival of cancer cells. Additionally, we indicate that targeting particular stages of cAMP-dependent processes might provide promising therapeutic opportunities for the effective management of patients with ovarian cancer.
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Affiliation(s)
- Agnieszka Kilanowska
- Department of Anatomy and Histology, Collegium Medicum, University of Zielona Gora, 65-046 Zielona Gora, Poland
- Correspondence: ; Tel.: +48-683-283-148
| | - Agnieszka Ziółkowska
- Department of Anatomy and Histology, Collegium Medicum, University of Zielona Gora, 65-046 Zielona Gora, Poland
| | - Piotr Stasiak
- Department of Anatomy and Histology, Collegium Medicum, University of Zielona Gora, 65-046 Zielona Gora, Poland
| | - Magdalena Gibas-Dorna
- Department of Anatomy and Histology, Collegium Medicum, University of Zielona Gora, 65-046 Zielona Gora, Poland
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Liu C, Guo X, Si H, Li G. A mink (Neovison vison) model of self-injury: Effects of CBP-CREB axis on neuronal damage and behavior. Front Vet Sci 2022; 9:975112. [DOI: 10.3389/fvets.2022.975112] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2022] [Accepted: 10/17/2022] [Indexed: 11/11/2022] Open
Abstract
ObjectiveSelf-injurious behavior (SIB) is a clinically challenging problem in the general population and several clinical disorders. However, the precise molecular mechanism of SIB is still not clear. In this paper, the systematic investigation of the genesis and development of SIB is conducted based on behavioral and pathophysiology studies in mink (Neovison vison) models.MethodThe night-vision video was used to observe the mink behavior, and the duration was a month. HE stain was performed to characterize the pathology change in the brain of a mink. IHC assay was performed to conduct the protein level detection of Iba-1, p-CREB, CBP, and p300 in the brain tissues. Elisa assay was used to examine the levels of NfL and NfH in serum and CSF of mink. The qRT-PCR assay was used to detect the expression of Bcl-2, NOR1, FoxO4, c-FOS, CBP, and p300 in brain tissues. Western blot was used to detect the protein levels of p-CREB, CBP, and p300 in brain tissues. We also used Evans Blue as a tracer to detect whether the blood-brain barrier was impaired in the brain of mink.ResultThe behavioral test, histopathological and molecular biology experiments were combined in this paper, and the results showed that CBP was related to SIB. Mechanism analysis showed that the dysregulation of CBP in brain-activated CREB signaling will result in nerve damage of the brain and SIB symptoms in minks. More importantly, the CBP-CREB interaction inhibitor might help relieve SIB and nerve damage in brain tissues.ConclusionOur results illustrate that the induction of CBP and the activation of CREB are novel mechanisms in the genesis of SIB. This finding indicates that the CBP-CREB axis is critical for SIB and demonstrates the efficacy of the CBP-CREB interaction inhibitor in treating these behaviors.
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Najih M, Nguyen HT, Martin LJ. Involvement of calmodulin-dependent protein kinase I in the regulation of the expression of connexin 43 in MA-10 tumor Leydig cells. Mol Cell Biochem 2022; 478:791-805. [PMID: 36094721 DOI: 10.1007/s11010-022-04553-7] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/12/2021] [Accepted: 08/30/2022] [Indexed: 11/30/2022]
Abstract
Connexin 43 (Cx43, also known as Gja1) is the most abundant testicular gap junction protein. It has a crucial role in the support of spermatogenesis by Sertoli cells in the seminiferous tubules as well as in androgen synthesis by Leydig cells. The multifunctional family of Ca2+/calmodulin-dependent protein kinases (CaMK) is composed of CaMK I, II, and IV and each can serve as a mediator of nuclear Ca2+ signals. These kinases can control gene expression by phosphorylation of key regulatory sites on transcription factors. Among these, AP-1 members cFos and cJun are interesting candidates that seem to cooperate with CaMKs to regulate Cx43 expression in Leydig cells. In this study, the Cx43 promoter region important for CaMK-dependent activation is characterized using co-transfection of plasmid reporter-constructs with different plasmids coding for CaMKs and/or AP-1 members in MA-10 Leydig cells. Here we report that the activation of Cx43 expression by cFos and cJun is increased by CaMKI. Furthermore, results from chromatin immunoprecipitation suggest that the recruitment of AP-1 family members to the proximal region of the Cx43 promoter may involve another uncharacterized AP-1 DNA regulatory element and/or protein-protein interactions with other partners. Thus, our data provide new insights into the molecular regulatory mechanisms that control mouse Cx43 transcription in testicular Leydig cells.
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Affiliation(s)
- Mustapha Najih
- Biology Department, Université de Moncton, 18, avenue Antonine Maillet, Moncton, NB, E1A 3E9, Canada
| | - Ha Tuyen Nguyen
- Biology Department, Université de Moncton, 18, avenue Antonine Maillet, Moncton, NB, E1A 3E9, Canada
| | - Luc J Martin
- Biology Department, Université de Moncton, 18, avenue Antonine Maillet, Moncton, NB, E1A 3E9, Canada.
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Activation of ADRB2/PKA Signaling Pathway Facilitates Lipid Synthesis in Meibocytes, and Beta-Blocker Glaucoma Drug Impedes PKA-Induced Lipid Synthesis by Inhibiting ADRB2. Int J Mol Sci 2022; 23:ijms23169478. [PMID: 36012741 PMCID: PMC9409328 DOI: 10.3390/ijms23169478] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2022] [Revised: 08/16/2022] [Accepted: 08/17/2022] [Indexed: 11/18/2022] Open
Abstract
Meibomian gland dysfunction is one of the main causes of dry eye disease and has limited therapeutic options. In this study, we investigated the biological function of the beta 2-adrenergic receptor (ADRB2)/protein kinase A (PKA) pathway in lipid synthesis and its underlying mechanisms in human meibomian gland epithelial cells (HMGECs). HMGECs were cultured in differentiation media with or without forskolin (an activator of adenylate cyclase), salbutamol (an ADRB2 agonist), or timolol (an ADRB2 antagonist) for up to 4 days. The phosphorylation of the cAMP-response element-binding protein (CREB) and the expression of peroxisome proliferator activator receptor (PPAR)γ and sterol regulatory element-binding protein (SREBP)-1 were measured by immunoblotting and quantitative PCR. Lipid synthesis was examined by LipidTOX immunostaining, AdipoRed assay, and Oil Red O staining. PKA pathway activation enhanced PPARγ expression and lipid synthesis in differentiated HMGECs. When treated with agonists of ADBR2 (upstream of the PKA signaling system), PPARγ expression and lipid synthesis were enhanced in HMGECs. The ADRB2 antagonist timolol showed the opposite effect. The activation of the ADRB2/PKA signaling pathway enhances lipid synthesis in HMGECs. These results provide a potential mechanism and therapeutic target for meibomian gland dysfunction, particularly in cases induced by beta-blocker glaucoma drugs.
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Ali M, LaCanna R, Lian Z, Huang J, Tan Y, Shao W, Yu X, Tian Y. Transcriptional responses to injury of regenerative lung alveolar epithelium. iScience 2022; 25:104843. [PMID: 35996586 PMCID: PMC9391595 DOI: 10.1016/j.isci.2022.104843] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2021] [Revised: 02/01/2022] [Accepted: 07/22/2022] [Indexed: 11/23/2022] Open
Abstract
The significance of alveolar epithelial type 2 (AT2) cell proliferation for lung alveolar epithelial homeostasis and regeneration after injury has been widely accepted. However, the heterogeneity of AT2 cell population for cell proliferation capacity remains disputed. By single-cell RNA sequencing and genetic lineage labeling using the Ki67 knock-in mouse model, we map all proliferative AT2 cells in homeostatic and regenerating murine lungs after injury induced by Streptococcus pneumoniae infection. The proliferative AT2 cell population displays a unique transcriptional program, which is regulated by activating transcription factor 3 (ATF3) and thyroid hormone receptor alpha (THRA) transcription factors. Overexpression of these two transcription factors in AT2 cells promoted AT2 cell proliferation and improved lung function after injury. These results indicate that increased expression of ATF3 and THRA at the onset of lung epithelial regeneration is required to permit rapid AT2 cell proliferation and hence progression through the recovery of lung epithelium.
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Affiliation(s)
- Mir Ali
- Department of Cardiovascular Sciences, Center for Translational Medicine, Temple University Lewis Katz School of Medicine, Philadelphia, PA 19140, USA
| | - Ryan LaCanna
- Department of Cardiovascular Sciences, Center for Translational Medicine, Temple University Lewis Katz School of Medicine, Philadelphia, PA 19140, USA
| | - Zhaorui Lian
- Coriell Institute for Medical Research, Camden, NJ 08103, USA
| | - Jian Huang
- Coriell Institute for Medical Research, Camden, NJ 08103, USA
| | - Yinfei Tan
- Fox Chase Cancer Center, Temple University Health System, Philadelphia, PA 19111, USA
| | - Wenna Shao
- Joint International Research Laboratory of Metabolic and Developmental Sciences, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
| | - Xiang Yu
- Joint International Research Laboratory of Metabolic and Developmental Sciences, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China
| | - Ying Tian
- Department of Cardiovascular Sciences, Center for Translational Medicine, Temple University Lewis Katz School of Medicine, Philadelphia, PA 19140, USA
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Shrestha J, Santerre M, Allen CNS, Arjona SP, Merali C, Mukerjee R, Chitrala KN, Park J, Bagashev A, Bui V, Eugenin EA, Merali S, Kaul M, Chin J, Sawaya BE. HIV-1 gp120 Impairs Spatial Memory Through Cyclic AMP Response Element-Binding Protein. Front Aging Neurosci 2022; 14:811481. [PMID: 35615594 PMCID: PMC9124804 DOI: 10.3389/fnagi.2022.811481] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2021] [Accepted: 03/31/2022] [Indexed: 11/13/2022] Open
Abstract
HIV-associated neurocognitive disorders (HAND) remain an unsolved problem that persists despite using antiretroviral therapy. We have obtained data showing that HIV-gp120 protein contributes to neurodegeneration through metabolic reprogramming. This led to decreased ATP levels, lower mitochondrial DNA copy numbers, and loss of mitochondria cristae, all-important for mitochondrial biogenesis. gp120 protein also disrupted mitochondrial movement and synaptic plasticity. Searching for the mechanisms involved, we found that gp120 alters the cyclic AMP response element-binding protein (CREB) phosphorylation on serine residue 133 necessary for its function as a transcription factor. Since CREB regulates the promoters of PGC1α and BDNF genes, we found that CREB dephosphorylation causes PGC1α and BDNF loss of functions. The data was validated in vitro and in vivo. The negative effect of gp120 was alleviated in cells and animals in the presence of rolipram, an inhibitor of phosphodiesterase protein 4 (PDE4), restoring CREB phosphorylation. We concluded that HIV-gp120 protein contributes to HAND via inhibition of CREB protein function.
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Affiliation(s)
- Jenny Shrestha
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
| | - Maryline Santerre
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
| | - Charles N. S. Allen
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
| | - Sterling P. Arjona
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
| | - Carmen Merali
- Department of Pharmaceutical Sciences, School of Pharmacy, Temple University, Philadelphia, PA, United States
| | - Ruma Mukerjee
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
| | | | - Jin Park
- Memory and Brain Research Center, Department of Neuroscience, Baylor College of Medicine, Houston, TX, United States
| | - Asen Bagashev
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
| | - Viet Bui
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
| | - Eliseo A. Eugenin
- Department of Neuroscience, Cell Biology, and Anatomy, The University of Texas Medical Branch, Galveston, TX, United States
| | - Salim Merali
- Department of Pharmaceutical Sciences, School of Pharmacy, Temple University, Philadelphia, PA, United States
| | - Marcus Kaul
- Infectious and Inflammatory Disease Center, Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA, United States
- Department of Psychiatry, University of California, San Diego, San Diego, CA, United States
- Division of Biomedical Sciences, School of Medicine, University of California, Riverside, Riverside, CA, United States
| | - Jeannie Chin
- Memory and Brain Research Center, Department of Neuroscience, Baylor College of Medicine, Houston, TX, United States
| | - Bassel E. Sawaya
- Molecular Studies of Neurodegenerative Diseases Lab, Philadelphia, PA, United States
- Fels Cancer Institute for Personalized Medicine Institute, Philadelphia, PA, United States
- Department of Neurology, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
- Department of Cancer and Cell Biology, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
- Department of Neural Sciences, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
- *Correspondence: Bassel E. Sawaya,
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Yin S, Song R, Ma J, Liu C, Wu Z, Cao G, Liu J, Zhang G, Zhang H, Sun R, Chen A, Wang Y. Receptor activity-modifying protein 1 regulates mouse skin fibroblast proliferation via the Gαi3-PKA-CREB-YAP axis. Cell Commun Signal 2022; 20:52. [PMID: 35413847 PMCID: PMC9004193 DOI: 10.1186/s12964-022-00852-0] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/09/2021] [Accepted: 03/01/2022] [Indexed: 01/13/2023] Open
Abstract
Background Skin innervation is crucial for normal wound healing. However, the relationship between nerve receptors and wound healing and the intrinsic mechanism remains to be further identified. In this study, we investigated the role of a calcitonin gene-related peptide (CGRP) receptor component, receptor activity‐modifying protein 1 (RAMP1), in mouse skin fibroblast (MSF) proliferation. Methods In vivo, Western blotting and immunohistochemical (IHC) staining of mouse skin wounds tissue was used to detect changes in RAMP1 expression. In vitro, RAMP1 was overexpressed in MSF cell lines by infection with Tet-On-Flag-RAMP1 lentivirus and doxycycline (DOX) induction. An IncuCyte S3 Live-Cell Analysis System was used to assess and compare the proliferation rate differences between different treatment groups. Total protein and subcellular extraction Western blot analysis, quantitative real-time-polymerase chain reaction (qPCR) analysis, and immunofluorescence (IF) staining analysis were conducted to detect signalling molecule expression and/or distribution. The CUT & RUN assay and dual-luciferase reporter assay were applied to measure protein-DNA interactions. Results RAMP1 expression levels were altered during skin wound healing in mice. RAMP1 overexpression promoted MSF proliferation. Mechanistically, total Yes-associated protein (YAP) and nuclear YAP protein expression was increased in RAMP1-overexpressing MSFs. RAMP1 overexpression increased inhibitory guanine nucleotide-binding protein (G protein) α subunit 3 (Gαi3) expression and activated downstream protein kinase A (PKA), and both elevated the expression of cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB) and activated it, promoting the transcription of YAP, elevating the total YAP level and promoting MSF proliferation. Conclusions Based on these data, we report, for the first time, that changes in the total RAMP1 levels during wound healing and RAMP1 overexpression alone can promote MSF proliferation via the Gαi3-PKA-CREB-YAP axis, a finding critical for understanding RAMP1 function, suggesting that this pathway is an attractive and accurate nerve target for skin wound treatment.
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