|
1
|
Cheng E, Lu R, Gerhold AR. Non-autonomous insulin signaling delays mitotic progression in C. elegans germline stem and progenitor cells. PLoS Genet 2024; 20:e1011351. [PMID: 39715269 PMCID: PMC11706408 DOI: 10.1371/journal.pgen.1011351] [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: 07/01/2024] [Revised: 01/07/2025] [Accepted: 11/25/2024] [Indexed: 12/25/2024] Open
Abstract
Stem and progenitor cell mitosis is essential for tissue development and homeostasis. How these cells ensure proper chromosome segregation, and thereby maintain mitotic fidelity, in the complex physiological environment of a living animal is poorly understood. Here we use in situ live-cell imaging of C. elegans germline stem and progenitor cells (GSPCs) to ask how the signaling environment influences stem and progenitor cell mitosis in vivo. Through a candidate screen we identify a new role for the insulin/IGF receptor (IGFR), daf-2, during GSPC mitosis. Mitosis is delayed in daf-2/IGFR mutants, and these delays require canonical, DAF-2/IGFR to DAF-16/FoxO insulin signaling, here acting cell non-autonomously from the soma. Interestingly, mitotic delays in daf-2/IGFR mutants depend on the spindle assembly checkpoint but are not accompanied by a loss of mitotic fidelity. Correspondingly, we show that caloric restriction, which delays GSPC mitosis and compromises mitotic fidelity, does not act via the canonical insulin signaling pathway, and instead requires AMP-activated kinase (AMPK). Together this work demonstrates that GSPC mitosis is influenced by at least two genetically separable signaling pathways and highlights the importance of signaling networks for proper stem and progenitor cell mitosis in vivo.
Collapse
Affiliation(s)
- Eric Cheng
- Department of Biology, McGill University, Montréal, Canada
| | - Ran Lu
- Department of Biology, McGill University, Montréal, Canada
| | | |
Collapse
|
|
2
|
Beitz A, Teves J, Oakes C, Johnstone C, Wang N, Brickman JM, Galloway KE. Cells transit through a quiescent-like state to convert to neurons at high rates. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.11.22.624928. [PMID: 39651159 PMCID: PMC11623504 DOI: 10.1101/2024.11.22.624928] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/11/2024]
Abstract
While transcription factors (TFs) provide essential cues for directing and redirecting cell fate, TFs alone are insufficient to drive cells to adopt alternative fates. Rather, transcription factors rely on receptive cell states to induce novel identities. Cell state emerges from and is shaped by cellular history and the activity of diverse processes. Here, we define the cellular and molecular properties of a highly receptive state amenable to transcription factor-mediated direct conversion from fibroblasts to induced motor neurons. Using a well-defined model of direct conversion to a post-mitotic fate, we identify the highly proliferative, receptive state that transiently emerges during conversion. Through examining chromatin accessibility, histone marks, and nuclear features, we find that cells reprogram from a state characterized by global reductions in nuclear size and transcriptional activity. Supported by globally increased levels of H3K27me3, cells enter a quiescent-like state of reduced RNA metabolism and elevated expression of REST and p27, markers of quiescent neural stem cells. From this transient state, cells convert to neurons at high rates. Inhibition of Ezh2, the catalytic subunit of PRC2 that deposits H3K27me3, abolishes conversion. Our work offers a roadmap to identify global changes in cellular processes that define cells with different conversion potentials that may generalize to other cell-fate transitions. Highlights Proliferation drives cells to a compact nuclear state that is receptive to TF-mediated conversion.Increased receptivity to TFs corresponds to reduced nuclear volumes.Reprogrammable cells display global, genome-wide increases in H3K27me3.High levels of H3K27me3 support cells' transits through a state of altered RNA metabolism.Inhibition of Ezh2 increases nuclear size, reduces the expression of the quiescence marker p27.Acute inhibition of Ezh2 abolishes motor neuron conversion. One Sentence Summary Cells transit through a quiescent-like state characterized by global reductions in nuclear size and transcriptional activity to convert to neurons at high rates.
Collapse
|
|
3
|
Cao W, Fan Q, Amparado G, Begic D, Godini R, Gopal S, Pocock R. A nucleic acid binding protein map of germline regulation in Caenorhabditis elegans. Nat Commun 2024; 15:6884. [PMID: 39128930 PMCID: PMC11317507 DOI: 10.1038/s41467-024-51212-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2024] [Accepted: 08/01/2024] [Indexed: 08/13/2024] Open
Abstract
Fertility requires the faithful proliferation of germ cells and their differentiation into gametes. Controlling these cellular states demands precise timing and expression of gene networks. Nucleic acid binding proteins (NBPs) play critical roles in gene expression networks that influence germ cell development. There has, however, been no functional analysis of the entire NBP repertoire in controlling in vivo germ cell development. Here, we analyzed germ cell states and germline architecture to systematically investigate the function of 364 germline-expressed NBPs in the Caenorhabditis elegans germ line. Using germline-specific knockdown, automated germ cell counting, and high-content analysis of germ cell nuclei and plasma membrane organization, we identify 156 NBPs with discrete autonomous germline functions. By identifying NBPs that control the germ cell cycle, proliferation, differentiation, germline structure and fertility, we have created an atlas for mechanistic dissection of germ cell behavior and gamete production.
Collapse
Affiliation(s)
- Wei Cao
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia.
| | - Qi Fan
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia
| | - Gemmarie Amparado
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia
| | - Dean Begic
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia
| | - Rasoul Godini
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia
| | - Sandeep Gopal
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia.
- Lund Stem Cell Center, Department of Experimental Medical Science, Lund University, Lund, Sweden.
| | - Roger Pocock
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, 3800, Australia.
| |
Collapse
|
|
4
|
Amran A, Pigatto L, Farley J, Godini R, Pocock R, Gopal S. The matrisome landscape controlling in vivo germ cell fates. Nat Commun 2024; 15:4200. [PMID: 38760342 PMCID: PMC11101451 DOI: 10.1038/s41467-024-48283-4] [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: 08/17/2023] [Accepted: 04/26/2024] [Indexed: 05/19/2024] Open
Abstract
The developmental fate of cells is regulated by intrinsic factors and the extracellular environment. The extracellular matrix (matrisome) delivers chemical and mechanical cues that can modify cellular development. However, comprehensive understanding of how matrisome factors control cells in vivo is lacking. Here we show that specific matrisome factors act individually and collectively to control germ cell development. Surveying development of undifferentiated germline stem cells through to mature oocytes in the Caenorhabditis elegans germ line enabled holistic functional analysis of 443 conserved matrisome-coding genes. Using high-content imaging, 3D reconstruction, and cell behavior analysis, we identify 321 matrisome genes that impact germ cell development, the majority of which (>80%) are undescribed. Our analysis identifies key matrisome networks acting autonomously and non-autonomously to coordinate germ cell behavior. Further, our results demonstrate that germ cell development requires continual remodeling of the matrisome landscape. Together, this study provides a comprehensive platform for deciphering how extracellular signaling controls cellular development and anticipate this will establish new opportunities for manipulating cell fates.
Collapse
Affiliation(s)
- Aqilah Amran
- Department of Experimental Medical Science, Lund University, Lund, Sweden
- Lund Stem Cell Center, Lund University, Lund, Sweden
- Lund Cancer Center, Lund University, Lund, Sweden
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute. Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, Australia
| | - Lara Pigatto
- Department of Experimental Medical Science, Lund University, Lund, Sweden
- Lund Stem Cell Center, Lund University, Lund, Sweden
- Lund Cancer Center, Lund University, Lund, Sweden
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute. Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, Australia
| | - Johanna Farley
- Department of Experimental Medical Science, Lund University, Lund, Sweden
- Lund Stem Cell Center, Lund University, Lund, Sweden
- Lund Cancer Center, Lund University, Lund, Sweden
| | - Rasoul Godini
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute. Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, Australia
| | - Roger Pocock
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute. Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, Australia.
| | - Sandeep Gopal
- Department of Experimental Medical Science, Lund University, Lund, Sweden.
- Lund Stem Cell Center, Lund University, Lund, Sweden.
- Lund Cancer Center, Lund University, Lund, Sweden.
- Development and Stem Cells Program, Monash Biomedicine Discovery Institute. Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC, Australia.
| |
Collapse
|
|
5
|
Pires da Silva A, Kelleher R, Reynoldson L. Decoding lifespan secrets: the role of the gonad in Caenorhabditis elegans aging. FRONTIERS IN AGING 2024; 5:1380016. [PMID: 38605866 PMCID: PMC11008531 DOI: 10.3389/fragi.2024.1380016] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 01/31/2024] [Accepted: 03/18/2024] [Indexed: 04/13/2024]
Abstract
The gonad has become a central organ for understanding aging in C. elegans, as removing the proliferating stem cells in the germline results in significant lifespan extension. Similarly, when starvation in late larval stages leads to the quiescence of germline stem cells the adult nematode enters reproductive diapause, associated with an extended lifespan. This review summarizes recent advancements in identifying the mechanisms behind gonad-mediated lifespan extension, including comparisons with other nematodes and the role of lipid signaling and transcriptional changes. Given that the gonad also mediates lifespan regulation in other invertebrates and vertebrates, elucidating the underlying mechanisms may help to gain new insights into the mechanisms and evolution of aging.
Collapse
|
|
6
|
Bollen DP, Reddy KC, Lascarez-Lagunas LI, Kim DH, Colaiácovo MP. Germline mitotic quiescence and cell death are induced in Caenorhabditis elegans by exposure to pathogenic Pseudomonas aeruginosa. Genetics 2024; 226:iyad197. [PMID: 37956057 PMCID: PMC10763535 DOI: 10.1093/genetics/iyad197] [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: 08/08/2023] [Revised: 08/08/2023] [Accepted: 11/03/2023] [Indexed: 11/15/2023] Open
Abstract
The impact of exposure to microbial pathogens on animal reproductive capacity and germline physiology is not well understood. The nematode Caenorhabditis elegans is a bacterivore that encounters pathogenic microbes in its natural environment. How pathogenic bacteria affect host reproductive capacity of C. elegans is not well understood. Here, we show that exposure of C. elegans hermaphrodites to the Gram-negative pathogen Pseudomonas aeruginosa causes a marked reduction in brood size with concomitant reduction in the number of nuclei in the germline and gonad size. We define 2 processes that are induced that contribute to the decrease in the number of germ cell nuclei. First, we observe that infection with P. aeruginosa leads to the induction of germ cell apoptosis. Second, we observe that this exposure induces mitotic quiescence in the proliferative zone of the C. elegans gonad. Importantly, these processes appear to be reversible; when animals are removed from the presence of P. aeruginosa, germ cell apoptosis is abated, germ cell nuclei numbers increase, and brood sizes recover. The reversible germline dynamics during exposure to P. aeruginosa may represent an adaptive response to improve survival of progeny and may serve to facilitate resource allocation that promotes survival during pathogen infection.
Collapse
Affiliation(s)
- Daniel P Bollen
- Division of Infectious Diseases and Department of Pediatrics, Boston Children's Hospital and Harvard Medical School, Boston, MA 02115, USA
- Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
| | - Kirthi C Reddy
- Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
- Molecular Neurobiology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037, USA
| | | | - Dennis H Kim
- Division of Infectious Diseases and Department of Pediatrics, Boston Children's Hospital and Harvard Medical School, Boston, MA 02115, USA
| | - Monica P Colaiácovo
- Department of Genetics, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA
| |
Collapse
|
|
7
|
Bollen DP, Reddy KC, Kim DH, Colaiácovo MP. Germline mitotic quiescence and programmed cell death are induced in C. elegans by exposure to pathogenic P. aeruginosa. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.08.08.552522. [PMID: 37609207 PMCID: PMC10441368 DOI: 10.1101/2023.08.08.552522] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/24/2023]
Abstract
The impact of exposure to microbial pathogens on animal reproductive capacity and germline physiology is not well understood. The nematode Caenorhabditis elegans is a bacterivore that encounters pathogenic microbes in its natural environment. How pathogenic bacteria affect host reproductive capacity of C. elegans is not well understood. Here, we show that exposure of C. elegans hermaphrodites to the Gram-negative pathogen Pseudomonas aeruginosa causes a marked reduction in brood size with concomitant reduction in the number of nuclei in the germline and gonad size. We define two processes that are induced that contribute to the decrease in the number of germ cell nuclei. First, we observe that infection with P. aeruginosa leads to the induction of programmed germ cell death. Second, we observe that this exposure induces mitotic quiescence in the proliferative zone of the C. elegans gonad. Importantly, these processes appear to be reversible; when animals are removed from the presence of P. aeruginosa, germ cell death is abated, germ cell nuclei numbers increase, and brood sizes recover. The reversible germline dynamics during exposure to P. aeruginosa may represent an adaptive response to improve survival of progeny and may serve to facilitate resource allocation that promotes survival during pathogen infection.
Collapse
Affiliation(s)
- Daniel P. Bollen
- Division of Infectious Diseases and Department of Pediatrics, Boston Children’s Hospital and Harvard Medical School, Boston, MA 02115, USA
- Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
| | - Kirthi C. Reddy
- Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
| | - Dennis H. Kim
- Division of Infectious Diseases and Department of Pediatrics, Boston Children’s Hospital and Harvard Medical School, Boston, MA 02115, USA
| | | |
Collapse
|
|
8
|
Fausett SR, Sandjak A, Billard B, Braendle C. Higher-order epistasis shapes natural variation in germ stem cell niche activity. Nat Commun 2023; 14:2824. [PMID: 37198172 DOI: 10.1038/s41467-023-38527-0] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2022] [Accepted: 05/05/2023] [Indexed: 05/19/2023] Open
Abstract
To study how natural allelic variation explains quantitative developmental system variation, we characterized natural differences in germ stem cell niche activity, measured as progenitor zone (PZ) size, between two Caenorhabditis elegans isolates. Linkage mapping yielded candidate loci on chromosomes II and V, and we found that the isolate with a smaller PZ size harbours a 148 bp promoter deletion in the Notch ligand, lag-2/Delta, a central signal promoting germ stem cell fate. As predicted, introducing this deletion into the isolate with a large PZ resulted in a smaller PZ size. Unexpectedly, restoring the deleted ancestral sequence in the isolate with a smaller PZ did not increase-but instead further reduced-PZ size. These seemingly contradictory phenotypic effects are explained by epistatic interactions between the lag-2/Delta promoter, the chromosome II locus, and additional background loci. These results provide first insights into the quantitative genetic architecture regulating an animal stem cell system.
Collapse
Affiliation(s)
- Sarah R Fausett
- Université Côte d'Azur, CNRS, Inserm, IBV, Nice, France.
- Department of Biology and Marine Biology, University of North Carolina Wilmington, Wilmington, NC, USA.
| | - Asma Sandjak
- Université Côte d'Azur, CNRS, Inserm, IBV, Nice, France
| | | | | |
Collapse
|
|
9
|
Ferdous AS, Costa Dos Santos SJ, Kanzler CR, Shin H, Carrick BH, Crittenden SL, Wickens M, Kimble J. The in vivo functional significance of PUF hub partnerships in C. elegans germline stem cells. Development 2023; 150:dev201705. [PMID: 37070766 PMCID: PMC10259659 DOI: 10.1242/dev.201705] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2023] [Accepted: 03/29/2023] [Indexed: 04/19/2023]
Abstract
PUF RNA-binding proteins are conserved stem cell regulators. Four PUF proteins govern self-renewal of Caenorhabditis elegans germline stem cells together with two intrinsically disordered proteins, LST-1 and SYGL-1. Based on yeast two-hybrid results, we previously proposed a composite self-renewal hub in the stem cell regulatory network, with eight PUF partnerships and extensive redundancy. Here, we investigate LST-1-PUF and SYGL-1-PUF partnerships and their molecular activities in their natural context - nematode stem cells. We confirm LST-1-PUF partnerships and their specificity to self-renewal PUFs by co-immunoprecipitation and show that an LST-1(AmBm) mutant defective for PUF-interacting motifs does not complex with PUFs in nematodes. LST-1(AmBm) is used to explore the in vivo functional significance of the LST-1-PUF partnership. Tethered LST-1 requires this partnership to repress expression of a reporter RNA, and LST-1 requires the partnership to co-immunoprecipitate with NTL-1/Not1 of the CCR4-NOT complex. We suggest that the partnership provides multiple molecular interactions that work together to form an effector complex on PUF target RNAs in vivo. Comparison of LST-1-PUF and Nanos-Pumilio reveals fundamental molecular differences, making LST-1-PUF a distinct paradigm for PUF partnerships.
Collapse
Affiliation(s)
- Ahlan S. Ferdous
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | | | - Charlotte R. Kanzler
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Heaji Shin
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Brian H. Carrick
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Sarah L. Crittenden
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Marvin Wickens
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Judith Kimble
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| |
Collapse
|
|
10
|
Ferdous AS, Costa Dos Santos SJ, Kanzler CR, Shin H, Carrick BH, Crittenden SL, Wickens M, Kimble J. Functional significance of PUF partnerships in C. elegans germline stem cells. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.02.15.528708. [PMID: 36824876 PMCID: PMC9949348 DOI: 10.1101/2023.02.15.528708] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 04/26/2023]
Abstract
PUF RNA-binding proteins are conserved stem cell regulators. Four PUF proteins govern self-renewal of C. elegans germline stem cells together with two intrinsically disordered proteins, LST-1 and SYGL-1. Based on yeast two-hybrid results, we proposed a composite self-renewal hub in the stem cell regulatory network, with eight PUF partnerships and extensive redundancy. Here, we investigate LST-1-PUF and SYGL-1-PUF partnerships and their molecular activities in their natural context - nematode stem cells. We confirm LST-1-PUF partnerships and their specificity to self-renewal PUFs by co-immunoprecipitation and show that an LST-1(A m B m ) mutant defective for PUF-interacting motifs does not complex with PUFs in nematodes. LST-1(A m B m ) is used to explore the functional significance of the LST-1-PUF partnership. Tethered LST-1 requires the partnership to repress expression of a reporter RNA, and LST-1 requires the partnership to co-immunoprecipitate with NTL-1/Not1 of the CCR4-NOT complex. We suggest that the partnership provides multiple molecular interactions that work together to form an effector complex on PUF target RNAs. Comparison of PUF-LST-1 and Pumilio-Nanos reveals fundamental molecular differences, making PUF-LST-1 a distinct paradigm for PUF partnerships. Summary statement Partnerships between PUF RNA-binding proteins and intrinsically disordered proteins are essential for stem cell maintenance and RNA repression.
Collapse
Affiliation(s)
- Ahlan S Ferdous
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | | | - Charlotte R Kanzler
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Heaji Shin
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Brian H Carrick
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Sarah L Crittenden
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Marvin Wickens
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Judith Kimble
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| |
Collapse
|
|
11
|
Park Y, Gaddy M, Hyun M, Jones ME, Aslam HM, Lee MH. Genetic and Chemical Controls of Sperm Fate and Spermatocyte Dedifferentiation via PUF-8 and MPK-1 in Caenorhabditis elegans. Cells 2023; 12:cells12030434. [PMID: 36766775 PMCID: PMC9913519 DOI: 10.3390/cells12030434] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2022] [Revised: 01/21/2023] [Accepted: 01/24/2023] [Indexed: 02/03/2023] Open
Abstract
Using the nematode C. elegans germline as a model system, we previously reported that PUF-8 (a PUF RNA-binding protein) and LIP-1 (a dual-specificity phosphatase) repress sperm fate at 20 °C and the dedifferentiation of spermatocytes into mitotic cells (termed "spermatocyte dedifferentiation") at 25 °C. Thus, double mutants lacking both PUF-8 and LIP-1 produce excess sperm at 20 °C, and their spermatocytes return to mitotically dividing cells via dedifferentiation at 25 °C, resulting in germline tumors. To gain insight into the molecular competence for spermatocyte dedifferentiation, we compared the germline phenotypes of three mutant strains that produce excess sperm-fem-3(q20gf), puf-8(q725); fem-3(q20gf), and puf-8(q725); lip-1(zh15). Spermatocyte dedifferentiation was not observed in fem-3(q20gf) mutants, but it was more severe in puf-8(q725); lip-1(zh15) than in puf-8(q725); fem-3(q20gf) mutants. These results suggest that MPK-1 (the C. elegans ERK1/2 MAPK ortholog) activation in the absence of PUF-8 is required to promote spermatocyte dedifferentiation. This idea was confirmed using Resveratrol (RSV), a potential activator of MPK-1 and ERK1/2 in C. elegans and human cells, respectively. Notably, spermatocyte dedifferentiation was significantly enhanced by RSV treatment in the absence of PUF-8, and its effect was blocked by mpk-1 RNAi. We, therefore, conclude that PUF-8 and MPK-1 are essential regulators for spermatocyte dedifferentiation and tumorigenesis. Since these regulators are broadly conserved, we suggest that similar regulatory circuitry may control cellular dedifferentiation and tumorigenesis in other organisms, including humans.
Collapse
Affiliation(s)
- Youngyong Park
- Division of Hematology/Oncology, Department of Internal Medicine, Brody School of Medicine at East Carolina University, Greenville, NC 27834, USA
| | - Matthew Gaddy
- Division of Hematology/Oncology, Department of Internal Medicine, Brody School of Medicine at East Carolina University, Greenville, NC 27834, USA
| | - Moonjung Hyun
- Biological Resources Research Group, Bioenvironmental Science & Toxicology Division, Korea Institute of Toxicology, Jinju 52834, Gyeongsangnam-do, Republic of Korea
| | - Mariah E. Jones
- Division of Hematology/Oncology, Department of Internal Medicine, Brody School of Medicine at East Carolina University, Greenville, NC 27834, USA
| | - Hafiz M. Aslam
- Division of Hematology/Oncology, Department of Internal Medicine, Brody School of Medicine at East Carolina University, Greenville, NC 27834, USA
| | - Myon Hee Lee
- Division of Hematology/Oncology, Department of Internal Medicine, Brody School of Medicine at East Carolina University, Greenville, NC 27834, USA
- Department of Biology, East Carolina University, Greenville, NC 27858, USA
- Correspondence:
| |
Collapse
|
|
12
|
Crittenden SL, Seidel HS, Kimble J. Analysis of the C. elegans Germline Stem Cell Pool. Methods Mol Biol 2023; 2677:1-36. [PMID: 37464233 DOI: 10.1007/978-1-0716-3259-8_1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 07/20/2023]
Abstract
The Caenorhabditis elegans germline is an excellent model for studying the genetic and molecular regulation of stem cell self-renewal and progression of cells from a stem cell state to a differentiated state. The germline tissue is organized in an assembly line with the germline stem cell (GSC) pool at one end and differentiated gametes at the other. A simple mesenchymal niche caps the GSC pool and maintains GSCs in an undifferentiated state by signaling through the conserved Notch pathway. Notch signaling activates transcription of the key GSC regulators lst-1 and sygl-1 proteins in a gradient through the GSC pool. LST-1 and SYGL-1 proteins work with PUF RNA regulators in a self-renewal hub to maintain the GSC pool. In this chapter, we present methods for characterizing the C. elegans GSC pool and early stages of germ cell differentiation. The methods include examination of germlines in living and fixed worms, cell cycle analysis, and analysis of markers. We also discuss assays to separate mutant phenotypes that affect the stem cell vs. differentiation decision from those that affect germ cell processes more generally.
Collapse
Affiliation(s)
- Sarah L Crittenden
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI, USA.
| | - Hannah S Seidel
- Department of Biology, Eastern Michigan University, Ypsilanti, MI, USA
| | - Judith Kimble
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI, USA
| |
Collapse
|
|
13
|
Brenner JL, Jyo EM, Mohammad A, Fox P, Jones V, Mardis E, Schedl T, Maine EM. TRIM-NHL protein, NHL-2, modulates cell fate choices in the C. elegans germ line. Dev Biol 2022; 491:43-55. [PMID: 36063869 PMCID: PMC9922029 DOI: 10.1016/j.ydbio.2022.08.010] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2022] [Revised: 07/19/2022] [Accepted: 08/27/2022] [Indexed: 12/01/2022]
Abstract
Many tissues contain multipotent stem cells that are critical for maintaining tissue function. In Caenorhabditis elegans, germline stem cells allow gamete production to continue in adulthood. In the gonad, GLP-1/Notch signaling from the distal tip cell niche to neighboring germ cells activates a complex regulatory network to maintain a stem cell population. GLP-1/Notch signaling positively regulates production of LST-1 and SYGL-1 proteins that, in turn, interact with a set of PUF/FBF proteins to positively regulate the stem cell fate. We previously described sog (suppressor of glp-1 loss of function) and teg (tumorous enhancer of glp-1 gain of function) genes that limit the stem cell fate and/or promote the meiotic fate. Here, we show that sog-10 is allelic to nhl-2. NHL-2 is a member of the conserved TRIM-NHL protein family whose members can bind RNA and ubiquitinate protein substrates. We show that NHL-2 acts, at least in part, by inhibiting the expression of PUF-3 and PUF-11 translational repressor proteins that promote the stem cell fate. Two other negative regulators of stem cell fate, CGH-1 (conserved germline helicase) and ALG-5 (Argonaute protein), may work with NHL-2 to modulate the stem cell population. In addition, NHL-2 activity promotes the male germ cell fate in XX animals.
Collapse
Affiliation(s)
- John L Brenner
- Department of Genetics, Washington University School of Medicine, St. Louis, MO, 63110, USA
| | - Erin M Jyo
- Department of Biology, Syracuse University, Syracuse, NY, 13210, USA
| | - Ariz Mohammad
- Department of Genetics, Washington University School of Medicine, St. Louis, MO, 63110, USA
| | - Paul Fox
- Department of Genetics, Washington University School of Medicine, St. Louis, MO, 63110, USA
| | - Vovanti Jones
- Department of Genetics, Washington University School of Medicine, St. Louis, MO, 63110, USA
| | - Elaine Mardis
- McDonnell Genome Institute, Washington University School of Medicine, St. Louis, MO, 63110, USA
| | - Tim Schedl
- Department of Genetics, Washington University School of Medicine, St. Louis, MO, 63110, USA.
| | - Eleanor M Maine
- Department of Biology, Syracuse University, Syracuse, NY, 13210, USA.
| |
Collapse
|
|
14
|
Mechanisms of germ cell survival and plasticity in Caenorhabditis elegans. Biochem Soc Trans 2022; 50:1517-1526. [PMID: 36196981 PMCID: PMC9704514 DOI: 10.1042/bst20220878] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2022] [Revised: 09/20/2022] [Accepted: 09/21/2022] [Indexed: 11/17/2022]
Abstract
Animals constantly encounter environmental and physiological stressors that threaten survival and fertility. Somatic stress responses and germ cell arrest/repair mechanisms are employed to withstand such challenges. The Caenorhabditis elegans germline combats stress by initiating mitotic germ cell quiescence to preserve genome integrity, and by removing meiotic germ cells to prevent inheritance of damaged DNA or to tolerate lack of germline nutrient supply. Here, we review examples of germline recovery from distinct stressors - acute starvation and defective splicing - where quiescent mitotic germ cells resume proliferation to repopulate a germ line following apoptotic removal of meiotic germ cells. These protective mechanisms reveal the plastic nature of germline stem cells.
Collapse
|
|
15
|
Cao W, Tran C, Archer SK, Gopal S, Pocock R. Functional recovery of the germ line following splicing collapse. Cell Death Differ 2022; 29:772-787. [PMID: 34663906 PMCID: PMC8991207 DOI: 10.1038/s41418-021-00891-z] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2021] [Revised: 09/30/2021] [Accepted: 10/04/2021] [Indexed: 11/09/2022] Open
Abstract
Splicing introns from precursor-messenger RNA (pre-mRNA) transcripts is essential for translating functional proteins. Here, we report that the previously uncharacterized Caenorhabditis elegans protein MOG-7 acts as a pre-mRNA splicing factor. Depleting MOG-7 from the C. elegans germ line causes intron retention in most germline-expressed genes, impeding the germ cell cycle, and causing defects in nuclear morphology, germ cell identity and sterility. Despite the deleterious consequences caused by MOG-7 loss, the adult germ line can functionally recover to produce viable and fertile progeny when MOG-7 is restored. Germline recovery is dependent on a burst of apoptosis that likely clears defective germ cells, and viable gametes generated from the proliferation of germ cells in the progenitor zone. Together, these findings reveal that MOG-7 is essential for germ cell development, and that the germ line can functionally recover after a collapse in RNA splicing.
Collapse
Affiliation(s)
- Wei Cao
- grid.1002.30000 0004 1936 7857Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC 3800 Australia
| | - Christopher Tran
- grid.1002.30000 0004 1936 7857Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC 3800 Australia
| | - Stuart K. Archer
- grid.1002.30000 0004 1936 7857Monash Bioinformatics Platform, Monash University, Melbourne, VIC 3800 Australia
| | - Sandeep Gopal
- grid.1002.30000 0004 1936 7857Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC 3800 Australia
| | - Roger Pocock
- grid.1002.30000 0004 1936 7857Development and Stem Cells Program, Monash Biomedicine Discovery Institute and Department of Anatomy and Developmental Biology, Monash University, Melbourne, VIC 3800 Australia
| |
Collapse
|
|
16
|
Mallick A, Jhaveri N, Jeon J, Chang Y, Shah K, Hosein H, Gupta BP. Genetic analysis of Caenorhabditis elegans pry-1/Axin suppressors identifies genes involved in reproductive structure development, stress responses, and aging. G3 GENES|GENOMES|GENETICS 2022; 12:6462200. [PMID: 35100345 PMCID: PMC9210326 DOI: 10.1093/g3journal/jkab430] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/17/2021] [Accepted: 11/22/2021] [Indexed: 01/15/2023]
Abstract
Abstract
The Axin family of scaffolding proteins regulates a wide array of developmental and post-developmental processes in eukaryotes. Studies in the nematode Caenorhabditis elegans have shown that the Axin homolog PRY-1 plays essential roles in multiple tissues. To understand the genetic network of pry-1, we focused on a set of genes that are differentially expressed in the pry-1-mutant transcriptome and are linked to reproductive structure development. Knocking down eight of the genes (spp-1, clsp-1, ard-1, rpn-7, cpz-1, his-7, cdk-1, and rnr-1) via RNA interference efficiently suppressed the multivulva phenotype of pry-1 mutants. In all cases, the ectopic induction of P3.p vulval precursor cell was also inhibited. The suppressor genes are members of known gene families in eukaryotes and perform essential functions. Our genetic interaction experiments revealed that in addition to their role in vulval development, these genes participate in one or more pry-1-mediated biological events. Whereas four of them (cpz-1, his-7, cdk-1, and rnr-1) function in both stress response and aging, two (spp-1 and ard-1) are specific to stress response. Altogether, these findings demonstrate the important role of pry-1 suppressors in regulating developmental and post-developmental processes in C. elegans. Given that the genes described in this study are conserved, future investigations of their interactions with Axin and their functional specificity promises to uncover the genetic network of Axin in metazoans.
Collapse
Affiliation(s)
- Avijit Mallick
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| | - Nikita Jhaveri
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| | - Jihae Jeon
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| | - Yvonne Chang
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| | - Krupali Shah
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| | - Hannah Hosein
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| | - Bhagwati P Gupta
- Department of Biology, McMaster University, Hamilton, ON L8S4K1, Canada
| |
Collapse
|
|
17
|
From primordial germ cells to spermatids in Caenorhabditis elegans. Semin Cell Dev Biol 2021; 127:110-120. [PMID: 34930663 DOI: 10.1016/j.semcdb.2021.12.005] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2021] [Revised: 11/17/2021] [Accepted: 12/07/2021] [Indexed: 12/24/2022]
Abstract
Development of a syncytial germline for gamete formation requires complex regulation of cytokinesis and cytoplasmic remodeling. Recently, several uncovered cellular events have been investigated in the Caenorhabditis elegans (C. elegans) germline. In these cellular processes, the factors involved in contractility are highly conserved with those of mitosis and meiosis. However, the underlying regulatory mechanisms are far more complicated than previously thought, likely due to the single syncytial germline structure. In this review, we highlight how the proteins involved in contractility ensure faithful cell division in different cellular contexts and how they contribute to maintaining intercellular bridge stability. In addition, we discuss the current understanding of the cellular events of cytokinesis and cytoplasmic remodeling during the development of the C. elegans germline, including progenitor germ cells, germ cells, and spermatocytes. Comparisons are made with relevant systems in Drosophila melanogaster (D. melanogaster) and other animal models.
Collapse
|
|
18
|
Cahoon CK, Libuda DE. Conditional immobilization for live imaging Caenorhabditis elegans using auxin-dependent protein depletion. G3-GENES GENOMES GENETICS 2021; 11:6362942. [PMID: 34534266 PMCID: PMC8527506 DOI: 10.1093/g3journal/jkab310] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/25/2021] [Accepted: 08/19/2021] [Indexed: 11/12/2022]
Abstract
The visualization of biological processes using fluorescent proteins and dyes in living organisms has enabled numerous scientific discoveries. The nematode Caenorhabditis elegans is a widely used model organism for live imaging studies since the transparent nature of the worm enables imaging of nearly all tissues within a whole, intact animal. While current techniques are optimized to enable the immobilization of hermaphrodite worms for live imaging, many of these approaches fail to successfully restrain the smaller male worms. To enable live imaging of worms of both sexes, we developed a new genetic, conditional immobilization tool that uses the auxin-inducible degron (AID) system to immobilize both adult and larval hermaphrodite and male worms for live imaging. Based on chromosome location, mutant phenotype, and predicted germline consequence, we identified and AID-tagged three candidate genes (unc-18, unc-104, and unc-52). Strains with these AID-tagged genes were placed on auxin and tested for mobility and germline defects. Among the candidate genes, auxin-mediated depletion of UNC-18 caused significant immobilization of both hermaphrodite and male worms that was also partially reversible upon removal from auxin. Notably, we found that male worms require a higher concentration of auxin for a similar amount of immobilization as hermaphrodites, thereby suggesting a potential sex-specific difference in auxin absorption and/or processing. In both males and hermaphrodites, depletion of UNC-18 did not largely alter fertility, germline progression, nor meiotic recombination. Finally, we demonstrate that this new genetic tool can successfully immobilize both sexes enabling live imaging studies of sexually dimorphic features in C. elegans.
Collapse
Affiliation(s)
- Cori K Cahoon
- Department of Biology, Institute of Molecular Biology, University of Oregon, Eugene, OR 97403-1229, USA
| | - Diana E Libuda
- Department of Biology, Institute of Molecular Biology, University of Oregon, Eugene, OR 97403-1229, USA
| |
Collapse
|
|
19
|
Edwards SL, Erdenebat P, Morphis AC, Kumar L, Wang L, Chamera T, Georgescu C, Wren JD, Li J. Insulin/IGF-1 signaling and heat stress differentially regulate HSF1 activities in germline development. Cell Rep 2021; 36:109623. [PMID: 34469721 PMCID: PMC8442575 DOI: 10.1016/j.celrep.2021.109623] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2020] [Revised: 05/25/2021] [Accepted: 08/06/2021] [Indexed: 12/13/2022] Open
Abstract
Germline development is sensitive to nutrient availability and environmental perturbation. Heat shock transcription factor 1 (HSF1), a key transcription factor driving the cellular heat shock response (HSR), is also involved in gametogenesis. The precise function of HSF1 (HSF-1 in C. elegans) and its regulation in germline development are poorly understood. Using the auxin-inducible degron system in C. elegans, we uncovered a role of HSF-1 in progenitor cell proliferation and early meiosis and identified a compact but important transcriptional program of HSF-1 in germline development. Interestingly, heat stress only induces the canonical HSR in a subset of germ cells but impairs HSF-1 binding at its developmental targets. Conversely, insulin/insulin growth factor 1 (IGF-1) signaling dictates the requirement for HSF-1 in germline development and functions through repressing FOXO/DAF-16 in the soma to activate HSF-1 in germ cells. We propose that this non-cell-autonomous mechanism couples nutrient-sensing insulin/IGF-1 signaling to HSF-1 activation to support homeostasis in rapid germline growth.
Collapse
Affiliation(s)
- Stacey L Edwards
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Purevsuren Erdenebat
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Allison C Morphis
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Lalit Kumar
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Lai Wang
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Tomasz Chamera
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Constantin Georgescu
- Genes & Human Disease Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Jonathan D Wren
- Genes & Human Disease Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
| | - Jian Li
- Aging and Metabolism Research Program, Oklahoma Medical Research Foundation, Oklahoma City, OK, USA.
| |
Collapse
|
|
20
|
Tolkin T, Hubbard EJA. Germline Stem and Progenitor Cell Aging in C. elegans. Front Cell Dev Biol 2021; 9:699671. [PMID: 34307379 PMCID: PMC8297657 DOI: 10.3389/fcell.2021.699671] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2021] [Accepted: 06/07/2021] [Indexed: 11/13/2022] Open
Abstract
Like many animals and humans, reproduction in the nematode C. elegans declines with age. This decline is the cumulative result of age-related changes in several steps of germline function, many of which are highly accessible for experimental investigation in this short-lived model organism. Here we review recent work showing that a very early and major contributing step to reproductive decline is the depletion of the germline stem and progenitor cell pool. Since many cellular and molecular aspects of stem cell biology and aging are conserved across animals, understanding mechanisms of age-related decline of germline stem and progenitor cells in C. elegans has broad implications for aging stem cells, germline stem cells, and reproductive aging.
Collapse
Affiliation(s)
- Theadora Tolkin
- Department of Cell Biology, Skirball Institute of Biomolecular Medicine, NYU Grossman School of Medicine, New York, NY, United States
| | - E Jane Albert Hubbard
- Department of Cell Biology, Skirball Institute of Biomolecular Medicine, NYU Grossman School of Medicine, New York, NY, United States
| |
Collapse
|
|
21
|
Chen D, Li C, Zhao Y, Zhou J, Wang Q, Xie Y. Bioinformatics analysis for the identification of differentially expressed genes and related signaling pathways in H. pylori-CagA transfected gastric cancer cells. PeerJ 2021; 9:e11203. [PMID: 33954041 PMCID: PMC8053379 DOI: 10.7717/peerj.11203] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2020] [Accepted: 03/11/2021] [Indexed: 12/25/2022] Open
Abstract
Aim Helicobacter pylori cytotoxin-associated protein A (CagA) is an important virulence factor known to induce gastric cancer development. However, the cause and the underlying molecular events of CagA induction remain unclear. Here, we applied integrated bioinformatics to identify the key genes involved in the process of CagA-induced gastric epithelial cell inflammation and can ceration to comprehend the potential molecular mechanisms involved. Materials and Methods AGS cells were transected with pcDNA3.1 and pcDNA3.1::CagA for 24 h. The transfected cells were subjected to transcriptome sequencing to obtain the expressed genes. Differentially expressed genes (DEG) with adjusted P value < 0.05, — logFC —> 2 were screened, and the R package was applied for gene ontology (GO) enrichment and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The differential gene protein–protein interaction (PPI) network was constructed using the STRING Cytoscape application, which conducted visual analysis to create the key function networks and identify the key genes. Next, the Kaplan–Meier plotter survival analysis tool was employed to analyze the survival of the key genes derived from the PPI network. Further analysis of the key gene expressions in gastric cancer and normal tissues were performed based on The Cancer Genome Atlas (TCGA) database and RT-qPCR verification. Results After transfection of AGS cells, the cell morphology changes in a hummingbird shape and causes the level of CagA phosphorylation to increase. Transcriptomics identified 6882 DEG, of which 4052 were upregulated and 2830 were downregulated, among which q-value < 0.05, FC > 2, and FC under the condition of ≤2. Accordingly, 1062 DEG were screened, of which 594 were upregulated and 468 were downregulated. The DEG participated in a total of 151 biological processes, 56 cell components, and 40 molecular functions. The KEGG pathway analysis revealed that the DEG were involved in 21 pathways. The PPI network analysis revealed three highly interconnected clusters. In addition, 30 DEG with the highest degree were analyzed in the TCGA database. As a result, 12 DEG were found to be highly expressed in gastric cancer, while seven DEG were related to the poor prognosis of gastric cancer. RT-qPCR verification results showed that Helicobacter pylori CagA caused up-regulation of BPTF, caspase3, CDH1, CTNNB1, and POLR2A expression. Conclusion The current comprehensive analysis provides new insights for exploring the effect of CagA in human gastric cancer, which could help us understand the molecular mechanism underlying the occurrence and development of gastric cancer caused by Helicobacter pylori.
Collapse
Affiliation(s)
- Dingyu Chen
- Key Laboratory of Endemic and Ethnic Diseases , Ministry of Education, Guizhou Medical University, Guiyang, China
| | - Chao Li
- Key Laboratory of Endemic and Ethnic Diseases , Ministry of Education, Guizhou Medical University, Guiyang, China
| | - Yan Zhao
- Key Laboratory of Endemic and Ethnic Diseases , Ministry of Education, Guizhou Medical University, Guiyang, China
| | - Jianjiang Zhou
- Key Laboratory of Endemic and Ethnic Diseases , Ministry of Education, Guizhou Medical University, Guiyang, China
| | - Qinrong Wang
- Key Laboratory of Endemic and Ethnic Diseases , Ministry of Education, Guizhou Medical University, Guiyang, China
| | - Yuan Xie
- Key Laboratory of Endemic and Ethnic Diseases , Ministry of Education, Guizhou Medical University, Guiyang, China
| |
Collapse
|
|
22
|
Hefel A, Honda M, Cronin N, Harrell K, Patel P, Spies M, Smolikove S. RPA complexes in Caenorhabditis elegans meiosis; unique roles in replication, meiotic recombination and apoptosis. Nucleic Acids Res 2021; 49:2005-2026. [PMID: 33476370 PMCID: PMC7913698 DOI: 10.1093/nar/gkaa1293] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/10/2020] [Revised: 12/24/2020] [Accepted: 12/29/2020] [Indexed: 12/20/2022] Open
Abstract
Replication Protein A (RPA) is a critical complex that acts in replication and promotes homologous recombination by allowing recombinase recruitment to processed DSB ends. Most organisms possess three RPA subunits (RPA1, RPA2, RPA3) that form a trimeric complex critical for viability. The Caenorhabditis elegans genome encodes RPA-1, RPA-2 and an RPA-2 paralog RPA-4. In our analysis, we determined that RPA-2 is critical for germline replication and normal repair of meiotic DSBs. Interestingly, RPA-1 but not RPA-2 is essential for somatic replication, in contrast to other organisms that require both subunits. Six different hetero- and homodimeric complexes containing permutations of RPA-1, RPA-2 and RPA-4 can be detected in whole animal extracts. Our in vivo studies indicate that RPA-1/4 dimer is less abundant in the nucleus and its formation is inhibited by RPA-2. While RPA-4 does not participate in replication or recombination, we find that RPA-4 inhibits RAD-51 filament formation and promotes apoptosis of a subset of damaged nuclei. Altogether these findings point to sub-functionalization and antagonistic roles of RPA complexes in C. elegans.
Collapse
Affiliation(s)
- Adam Hefel
- Department of Biology, The University of Iowa, Iowa City, IA 52242, USA
| | - Masayoshi Honda
- Department of Biochemistry, The University of Iowa Carver College of Medicine, Iowa City, IA 52242, USA
| | - Nicholas Cronin
- Department of Biochemistry, The University of Iowa Carver College of Medicine, Iowa City, IA 52242, USA
| | - Kailey Harrell
- Department of Biology, The University of Iowa, Iowa City, IA 52242, USA
| | - Pooja Patel
- Department of Biology, The University of Iowa, Iowa City, IA 52242, USA
| | - Maria Spies
- Department of Biochemistry, The University of Iowa Carver College of Medicine, Iowa City, IA 52242, USA
| | - Sarit Smolikove
- Department of Biology, The University of Iowa, Iowa City, IA 52242, USA
| |
Collapse
|
|
23
|
Developmental plasticity and the response to nutrient stress in Caenorhabditis elegans. Dev Biol 2021; 475:265-276. [PMID: 33549550 DOI: 10.1016/j.ydbio.2021.01.015] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2020] [Revised: 12/24/2020] [Accepted: 01/29/2021] [Indexed: 11/23/2022]
Abstract
Developmental plasticity refers the ability of an organism to adapt to various environmental stressors, one of which is nutritional stress. Caenorhabditis elegans require various nutrients to successfully progress through all the larval stages to become a reproductive adult. If nutritional criteria are not satisfied, development can slow or completely arrest. In poor growth conditions, the animal can enter various diapause stages, depending on its developmental progress. In C. elegans, there are three well-characterized diapauses: the L1 arrest, the dauer diapause, and adult reproductive diapause, each associated with drastic changes in metabolism and germline development. At the centre of these changes is AMP-activated protein kinase (AMPK). AMPK is a metabolic regulator that maintains energy homeostasis, particularly during times of nutrient stress. Without AMPK, metabolism is disrupted during dauer, leading to the rapid consumption of lipid stores as well as misregulation of metabolic enzymes, leading to reduced survival. During the L1 arrest and dauer diapause, AMPK is responsible for ensuring germline quiescence by modifying the germline chromatin landscape to maintain germ cell integrity until conditions improve. Similar to classic hormonal signalling, small RNAs also play a critical role in regulating development and behaviour in a cell non-autonomous fashion. Thus, during the challenges associated with developmental plasticity, AMPK summons an army of signalling pathways to work collectively to preserve reproductive fitness during these periods of unprecedented uncertainty.
Collapse
|
|
24
|
Zellag RM, Zhao Y, Poupart V, Singh R, Labbé JC, Gerhold AR. CentTracker: a trainable, machine-learning-based tool for large-scale analyses of Caenorhabditis elegans germline stem cell mitosis. Mol Biol Cell 2021; 32:915-930. [PMID: 33502892 PMCID: PMC8108535 DOI: 10.1091/mbc.e20-11-0716] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Investigating the complex interactions between stem cells and their native environment requires an efficient means to image them in situ. Caenorhabditis elegans germline stem cells (GSCs) are distinctly accessible for intravital imaging; however, long-term image acquisition and analysis of dividing GSCs can be technically challenging. Here we present a systematic investigation into the technical factors impacting GSC physiology during live imaging and provide an optimized method for monitoring GSC mitosis under minimally disruptive conditions. We describe CentTracker, an automated and generalizable image analysis tool that uses machine learning to pair mitotic centrosomes and that can extract a variety of mitotic parameters rapidly from large-scale data sets. We employ CentTracker to assess a range of mitotic features in a large GSC data set. We observe spatial clustering of mitoses within the germline tissue but no evidence that subpopulations with distinct mitotic profiles exist within the stem cell pool. We further find biases in GSC spindle orientation relative to the germline’s distal–proximal axis and thus the niche. The technical and analytical tools provided herein pave the way for large-scale screening studies of multiple mitotic processes in GSCs dividing in situ, in an intact tissue, in a living animal, under seemingly physiological conditions.
Collapse
Affiliation(s)
- Réda M Zellag
- Department of Biology, McGill University, Montréal, QC H2A 1B1, Canada.,Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Succursale Centre-ville, Montréal, QC H3C 3J7, Canada
| | - Yifan Zhao
- Department of Biology, McGill University, Montréal, QC H2A 1B1, Canada.,Present address: Harvard-MIT Health Sciences and Technology, 77 Massachusetts Avenue, Cambridge, MA 02139
| | - Vincent Poupart
- Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Succursale Centre-ville, Montréal, QC H3C 3J7, Canada
| | - Ramya Singh
- Department of Biology, McGill University, Montréal, QC H2A 1B1, Canada.,Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Succursale Centre-ville, Montréal, QC H3C 3J7, Canada
| | - Jean-Claude Labbé
- Institute for Research in Immunology and Cancer (IRIC), Université de Montréal, Succursale Centre-ville, Montréal, QC H3C 3J7, Canada.,Department of Pathology and Cell Biology, Université de Montréal, Succursale Centre-ville, Montréal, QC H3C 3J7, Canada
| | - Abigail R Gerhold
- Department of Biology, McGill University, Montréal, QC H2A 1B1, Canada
| |
Collapse
|
|
25
|
Mata-Cabana A, Pérez-Nieto C, Olmedo M. Nutritional control of postembryonic development progression and arrest in Caenorhabditis elegans. ADVANCES IN GENETICS 2020; 107:33-87. [PMID: 33641748 DOI: 10.1016/bs.adgen.2020.11.002] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/11/2023]
Abstract
Developmental programs are under strict genetic control that favors robustness of the process. In order to guarantee the same outcome in different environmental situations, development is modulated by input pathways, which inform about external conditions. In the nematode Caenorhabditis elegans, the process of postembryonic development involves a series of stereotypic cell divisions, the progression of which is controlled by the nutritional status of the animal. C. elegans can arrest development at different larval stages, leading to cell arrest of the relevant divisions of the stage. This means that studying the nutritional control of development in C. elegans we can learn about the mechanisms controlling cell division in an in vivo model. In this work, we reviewed the current knowledge about the nutrient sensing pathways that control the progression or arrest of development in response to nutrient availability, with a special focus on the arrest at the L1 stage.
Collapse
Affiliation(s)
- Alejandro Mata-Cabana
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, Avd. Reina Mercedes, Sevilla, Spain
| | - Carmen Pérez-Nieto
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, Avd. Reina Mercedes, Sevilla, Spain
| | - María Olmedo
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, Avd. Reina Mercedes, Sevilla, Spain.
| |
Collapse
|
|
26
|
Baugh LR, Hu PJ. Starvation Responses Throughout the Caenorhabditiselegans Life Cycle. Genetics 2020; 216:837-878. [PMID: 33268389 PMCID: PMC7768255 DOI: 10.1534/genetics.120.303565] [Citation(s) in RCA: 80] [Impact Index Per Article: 16.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/22/2020] [Accepted: 08/17/2020] [Indexed: 02/07/2023] Open
Abstract
Caenorhabditis elegans survives on ephemeral food sources in the wild, and the species has a variety of adaptive responses to starvation. These features of its life history make the worm a powerful model for studying developmental, behavioral, and metabolic starvation responses. Starvation resistance is fundamental to life in the wild, and it is relevant to aging and common diseases such as cancer and diabetes. Worms respond to acute starvation at different times in the life cycle by arresting development and altering gene expression and metabolism. They also anticipate starvation during early larval development, engaging an alternative developmental program resulting in dauer diapause. By arresting development, these responses postpone growth and reproduction until feeding resumes. A common set of signaling pathways mediates systemic regulation of development in each context but with important distinctions. Several aspects of behavior, including feeding, foraging, taxis, egg laying, sleep, and associative learning, are also affected by starvation. A variety of conserved signaling, gene regulatory, and metabolic mechanisms support adaptation to starvation. Early life starvation can have persistent effects on adults and their descendants. With its short generation time, C. elegans is an ideal model for studying maternal provisioning, transgenerational epigenetic inheritance, and developmental origins of adult health and disease in humans. This review provides a comprehensive overview of starvation responses throughout the C. elegans life cycle.
Collapse
Affiliation(s)
- L Ryan Baugh
- Department of Biology, Center for Genomic and Computational Biology, Duke University, Durham, North Carolina 27708 and
| | - Patrick J Hu
- Departments of Medicine and Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232
| |
Collapse
|
|
27
|
Wang X, Ellenbecker M, Hickey B, Day NJ, Osterli E, Terzo M, Voronina E. Antagonistic control of Caenorhabditis elegans germline stem cell proliferation and differentiation by PUF proteins FBF-1 and FBF-2. eLife 2020; 9:52788. [PMID: 32804074 PMCID: PMC7467723 DOI: 10.7554/elife.52788] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2019] [Accepted: 08/14/2020] [Indexed: 02/07/2023] Open
Abstract
Stem cells support tissue maintenance, but the mechanisms that coordinate the rate of stem cell self-renewal with differentiation at a population level remain uncharacterized. We find that two PUF family RNA-binding proteins FBF-1 and FBF-2 have opposite effects on Caenorhabditis elegans germline stem cell dynamics: FBF-1 restricts the rate of meiotic entry, while FBF-2 promotes both cell division and meiotic entry rates. Antagonistic effects of FBFs are mediated by their distinct activities toward the shared set of target mRNAs, where FBF-1-mediated post-transcriptional control requires the activity of CCR4-NOT deadenylase, while FBF-2 is deadenylase-independent and might protect the targets from deadenylation. These regulatory differences depend on protein sequences outside of the conserved PUF family RNA-binding domain. We propose that the opposing FBF-1 and FBF-2 activities serve to modulate stem cell division rate simultaneously with the rate of meiotic entry.
Collapse
Affiliation(s)
- Xiaobo Wang
- Division of Biological Sciences, University of Montana, Missoula, United States
| | - Mary Ellenbecker
- Division of Biological Sciences, University of Montana, Missoula, United States
| | - Benjamin Hickey
- Division of Biological Sciences, University of Montana, Missoula, United States
| | - Nicholas J Day
- Division of Biological Sciences, University of Montana, Missoula, United States
| | - Emily Osterli
- Division of Biological Sciences, University of Montana, Missoula, United States
| | - Mikaya Terzo
- Division of Biological Sciences, University of Montana, Missoula, United States
| | - Ekaterina Voronina
- Division of Biological Sciences, University of Montana, Missoula, United States
| |
Collapse
|
|
28
|
Gordon K. Recent Advances in the Genetic, Anatomical, and Environmental Regulation of the C. elegans Germ Line Progenitor Zone. J Dev Biol 2020; 8:E14. [PMID: 32707774 PMCID: PMC7559772 DOI: 10.3390/jdb8030014] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2020] [Revised: 07/13/2020] [Accepted: 07/15/2020] [Indexed: 12/16/2022] Open
Abstract
The C. elegans germ line and its gonadal support cells are well studied from a developmental genetics standpoint and have revealed many foundational principles of stem cell niche biology. Among these are the observations that a niche-like cell supports a self-renewing stem cell population with multipotential, differentiating daughter cells. While genetic features that distinguish stem-like cells from their differentiating progeny have been defined, the mechanisms that structure these populations in the germ line have yet to be explained. The spatial restriction of Notch activation has emerged as an important genetic principle acting in the distal germ line. Synthesizing recent findings, I present a model in which the germ stem cell population of the C. elegans adult hermaphrodite can be recognized as two distinct anatomical and genetic populations. This review describes the recent progress that has been made in characterizing the undifferentiated germ cells and gonad anatomy, and presents open questions in the field and new directions for research to pursue.
Collapse
Affiliation(s)
- Kacy Gordon
- Department of Biology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA
| |
Collapse
|
|
29
|
Gordon KL, Zussman JW, Li X, Miller C, Sherwood DR. Stem cell niche exit in C. elegans via orientation and segregation of daughter cells by a cryptic cell outside the niche. eLife 2020; 9:e56383. [PMID: 32692313 PMCID: PMC7467730 DOI: 10.7554/elife.56383] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/25/2020] [Accepted: 07/17/2020] [Indexed: 12/17/2022] Open
Abstract
Stem cells reside in and rely upon their niche to maintain stemness but must balance self-renewal with the production of daughters that leave the niche to differentiate. We discovered a mechanism of stem cell niche exit in the canonical C. elegans distal tip cell (DTC) germ stem cell niche mediated by previously unobserved, thin, membranous protrusions of the adjacent somatic gonad cell pair (Sh1). A disproportionate number of germ cell divisions were observed at the DTC-Sh1 interface. Stem-like and differentiating cell fates segregated across this boundary. Spindles polarized, pairs of daughter cells oriented between the DTC and Sh1, and Sh1 grew over the Sh1-facing daughter. Impeding Sh1 growth by RNAi to cofilin and Arp2/3 perturbed the DTC-Sh1 interface, reduced germ cell proliferation, and shifted a differentiation marker. Because Sh1 membrane protrusions eluded detection for decades, it is possible that similar structures actively regulate niche exit in other systems.
Collapse
Affiliation(s)
- Kacy L Gordon
- Department of Biology, The University of North Carolina at Chapel HillChapel HillUnited States
| | - Jay W Zussman
- Department of Biology, Duke UniversityDurhamUnited States
| | - Xin Li
- Department of Biology, The University of North Carolina at Chapel HillChapel HillUnited States
| | - Camille Miller
- Department of Biology, The University of North Carolina at Chapel HillChapel HillUnited States
| | - David R Sherwood
- Department of Biology, Duke UniversityDurhamUnited States
- Regeneration Next, Duke UniversityDurhamUnited States
| |
Collapse
|
|
30
|
Wong C, Roy R. AMPK Regulates Developmental Plasticity through an Endogenous Small RNA Pathway in Caenorhabditis elegans. Int J Mol Sci 2020; 21:ijms21062238. [PMID: 32213851 PMCID: PMC7139869 DOI: 10.3390/ijms21062238] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/01/2020] [Revised: 03/18/2020] [Accepted: 03/22/2020] [Indexed: 01/19/2023] Open
Abstract
Caenorhabditis elegans larvae can undergo developmental arrest upon entry into the dauer stage in response to suboptimal growth conditions. Dauer larvae can exit this stage in replete conditions with no reproductive consequence. During this diapause stage, the metabolic regulator AMP-activated protein kinase (AMPK) ensures that the germ line becomes quiescent to maintain germ cell integrity. Animals that lack all AMPK signalling undergo germline hyperplasia upon entering dauer, while those that recover from this stage become sterile. Neuronal AMPK expression in otherwise AMPK-deficient animals is sufficient for germline quiescence and germ cell integrity and its effects are likely mediated through an endogenous small RNA pathway. Upon impairing small RNA biosynthesis, the post-dauer fertility is restored in AMPK mutants. These data suggest that AMPK may function in neurons to relay a message through small RNAs to the germ cells to alter their quiescence in the dauer stage, thus challenging the permeability of the Weismann barrier.
Collapse
|
|
31
|
Carranza-García E, Navarro RE. Insights Into the Hypometabolic Stage Caused by Prolonged Starvation in L4-Adult Caenorhabditis elegans Hermaphrodites. Front Cell Dev Biol 2020; 8:124. [PMID: 32211406 PMCID: PMC7057233 DOI: 10.3389/fcell.2020.00124] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2019] [Accepted: 02/12/2020] [Indexed: 11/24/2022] Open
Abstract
Animals alter their reproductive cycles in response to changing nutritional conditions, to ensure that offspring production only occurs under favorable circumstances. These adaptive strategies include reversible hypometabolic states of dormancy such as “arrest” and “diapause.” The free-living nematode Caenorhabditis elegans can arrest its life cycle during some larval stages without modifying its anatomy and physiology until conditions improve but it can also modify its morphological and physiological features to cope with harsh conditions and transition into diapause. The well-defined “dauer” diapause was described more than 40 years ago and has been the subject of comprehensive investigations. The existence of another hypometabolic state, termed adult reproductive diapause (ARD), has been debated after it was first described 10 years ago. Here, we review the current knowledge regarding the effect of food deprivation during the pre-reproductive larval and adult stages on overall organismal homeostasis, highlighting the implications on germ cell maintenance and fertility preservation.
Collapse
Affiliation(s)
- E Carranza-García
- Departamento de Biología Celular y Desarrollo, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Mexico City, Mexico
| | - Rosa E Navarro
- Departamento de Biología Celular y Desarrollo, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Mexico City, Mexico
| |
Collapse
|
|
32
|
Haupt KA, Law KT, Enright AL, Kanzler CR, Shin H, Wickens M, Kimble J. A PUF Hub Drives Self-Renewal in Caenorhabditis elegans Germline Stem Cells. Genetics 2020; 214:147-161. [PMID: 31740451 PMCID: PMC6944405 DOI: 10.1534/genetics.119.302772] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2019] [Accepted: 11/05/2019] [Indexed: 01/12/2023] Open
Abstract
Stem cell regulation relies on extrinsic signaling from a niche plus intrinsic factors that respond and drive self-renewal within stem cells. A priori, loss of niche signaling and loss of the intrinsic self-renewal factors might be expected to have equivalent stem cell defects. Yet this simple prediction has not been borne out for most stem cells, including Caenorhabditis elegans germline stem cells (GSCs). The central regulators of C. elegans GSCs include extrinsically acting GLP-1/Notch signaling from the niche; intrinsically acting RNA-binding proteins in the PUF family, termed FBF-1 and FBF-2 (collectively FBF); and intrinsically acting PUF partner proteins that are direct Notch targets. Abrogation of either GLP-1/Notch signaling or its targets yields an earlier and more severe GSC defect than loss of FBF-1 and FBF-2, suggesting that additional intrinsic regulators must exist. Here, we report that those missing regulators are two additional PUF proteins, PUF-3 and PUF-11 Remarkably, an fbf-1fbf-2 ; puf-3puf-11 quadruple null mutant has a GSC defect virtually identical to that of a glp-1/Notch null mutant. PUF-3 and PUF-11 both affect GSC maintenance, both are expressed in GSCs, and epistasis experiments place them at the same position as FBF within the network. Therefore, action of PUF-3 and PUF-11 explains the milder GSC defect in fbf-1fbf-2 mutants. We conclude that a "PUF hub," comprising four PUF proteins and two PUF partners, constitutes the intrinsic self-renewal node of the C. elegans GSC RNA regulatory network. Discovery of this hub underscores the significance of PUF RNA-binding proteins as key regulators of stem cell maintenance.
Collapse
Affiliation(s)
- Kimberly A Haupt
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| | - Kimberley T Law
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| | - Amy L Enright
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| | - Charlotte R Kanzler
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| | - Heaji Shin
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| | - Marvin Wickens
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| | - Judith Kimble
- Department of Biochemistry, University of Wisconsin-Madison, Wisconsin 53706
| |
Collapse
|
|
33
|
Hubbard EJA, Schedl T. Biology of the Caenorhabditis elegans Germline Stem Cell System. Genetics 2019; 213:1145-1188. [PMID: 31796552 PMCID: PMC6893382 DOI: 10.1534/genetics.119.300238] [Citation(s) in RCA: 86] [Impact Index Per Article: 14.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2019] [Accepted: 09/09/2019] [Indexed: 12/14/2022] Open
Abstract
Stem cell systems regulate tissue development and maintenance. The germline stem cell system is essential for animal reproduction, controlling both the timing and number of progeny through its influence on gamete production. In this review, we first draw general comparisons to stem cell systems in other organisms, and then present our current understanding of the germline stem cell system in Caenorhabditis elegans In contrast to stereotypic somatic development and cell number stasis of adult somatic cells in C. elegans, the germline stem cell system has a variable division pattern, and the system differs between larval development, early adult peak reproduction and age-related decline. We discuss the cell and developmental biology of the stem cell system and the Notch regulated genetic network that controls the key decision between the stem cell fate and meiotic development, as it occurs under optimal laboratory conditions in adult and larval stages. We then discuss alterations of the stem cell system in response to environmental perturbations and aging. A recurring distinction is between processes that control stem cell fate and those that control cell cycle regulation. C. elegans is a powerful model for understanding germline stem cells and stem cell biology.
Collapse
Affiliation(s)
- E Jane Albert Hubbard
- Skirball Institute of Biomolecular Medicine, Departments of Cell Biology and Pathology, New York University School of Medicine, New York 10016
| | - Tim Schedl
- and Department of Genetics, Washington University School of Medicine, St. Louis, Missouri 63110
| |
Collapse
|
|
34
|
Matthews H, Noulin F. Unexpected encounter of the parasitic kind. World J Stem Cells 2019; 11:904-919. [PMID: 31768219 PMCID: PMC6851008 DOI: 10.4252/wjsc.v11.i11.904] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/21/2019] [Revised: 08/10/2019] [Accepted: 09/13/2019] [Indexed: 02/06/2023] Open
Abstract
Both parasitology and stem cell research are important disciplines in their own right. Parasites are a real threat to human health causing a broad spectrum of diseases and significant annual rates morbidity and mortality globally. Stem cell research, on the other hand, focuses on the potential for regenerative medicine for a range of diseases including cancer and regenerative therapies. Though these two topics might appear distant, there are some “unexpected encounters”. In this review, we summarise the various links between parasites and stem cells. First, we discuss how parasites’ own stem cells represent interesting models of regeneration that can be translated to human stem cell regeneration. Second, we explore the interactions between parasites and host stem cells during the course of infection. Third, we investigate from a clinical perspective, how stem cell regeneration can be exploited to help circumvent the damage induced by parasitic infection and its potential to serve as treatment options for parasitic diseases in the future. Finally, we discuss the importance of screening for pathogens during organ transplantation by presenting some clinical cases of parasitic infection following stem cell therapy.
Collapse
Affiliation(s)
- Holly Matthews
- Centre for Applied Entomology and Parasitology, School of Life Sciences, Keele University, Keele ST5 5BG, United Kingdom
| | - Florian Noulin
- Centre for Applied Entomology and Parasitology, School of Life Sciences, Keele University, Keele ST5 5BG, United Kingdom
| |
Collapse
|
|
35
|
McKeown CR, Cline HT. Nutrient restriction causes reversible G2 arrest in Xenopus neural progenitors. Development 2019; 146:146/20/dev178871. [PMID: 31649012 DOI: 10.1242/dev.178871] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2019] [Accepted: 09/05/2019] [Indexed: 01/23/2023]
Abstract
Nutrient status affects brain development; however, the effects of nutrient availability on neural progenitor cell proliferation in vivo are poorly understood. Without food, Xenopus laevis tadpoles enter a period of stasis during which neural progenitor proliferation is drastically reduced, but resumes when food becomes available. Here, we investigate how neural progenitors halt cell division in response to nutrient restriction and subsequently re-enter the cell cycle upon feeding. We demonstrate that nutrient restriction causes neural progenitors to arrest in G2 of the cell cycle with increased DNA content, and that nutrient availability triggers progenitors to re-enter the cell cycle at M phase. Initiation of the nutrient restriction-induced G2 arrest is rapamycin insensitive, but cell cycle re-entry requires mTOR. Finally, we show that activation of insulin receptor signaling is sufficient to increase neural progenitor cell proliferation in the absence of food. A G2 arrest mechanism provides an adaptive strategy to control brain development in response to nutrient availability by triggering a synchronous burst of cell proliferation when nutrients become available. This may be a general cellular mechanism that allows developmental flexibility during times of limited resources.
Collapse
Affiliation(s)
| | - Hollis T Cline
- Department of Neuroscience, Scripps Research, La Jolla, CA 92037, USA
| |
Collapse
|
|
36
|
Haupt KA, Enright AL, Ferdous AS, Kershner AM, Shin H, Wickens M, Kimble J. The molecular basis of LST-1 self-renewal activity and its control of stem cell pool size. Development 2019; 146:dev.181644. [PMID: 31515205 DOI: 10.1242/dev.181644] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2019] [Accepted: 09/05/2019] [Indexed: 01/01/2023]
Abstract
PUF RNA-binding proteins have diverse roles in animal development, with a broadly conserved role in stem cells. Two paradigmatic PUF proteins, FBF-1 and FBF-2, promote both self-renewal and differentiation in the C. elegans germline. The LST-1 protein is a pivotal regulator of self-renewal and is oncogenic when mis-expressed. Here, we demonstrate that LST-1 self-renewal activity resides within a predicted disordered region that harbors two KXXL motifs. We find that the KXXL motifs mediate the binding of LST-1 to FBF, and that point mutations of these motifs abrogate LST-1 self-renewal activity. The LST-1-FBF partnership is therefore crucial to stem cell maintenance and is a key element in the FBF regulatory network. A distinct region within LST-1 determines its spatial expression and size of the GSC pool. Most importantly, the molecular understanding of how an IDR-rich protein works in an essential partnership with a conserved stem cell regulator and RNA-binding protein suggests broad new avenues for combinatorial control.
Collapse
Affiliation(s)
- Kimberly A Haupt
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Amy L Enright
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Ahlan S Ferdous
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Aaron M Kershner
- Howard Hughes Medical Institute, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Heaji Shin
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Marvin Wickens
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA
| | - Judith Kimble
- Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, USA .,Howard Hughes Medical Institute, University of Wisconsin-Madison, Madison, WI 53706, USA
| |
Collapse
|
|
37
|
Lara-Gonzalez P, Moyle MW, Budrewicz J, Mendoza-Lopez J, Oegema K, Desai A. The G2-to-M Transition Is Ensured by a Dual Mechanism that Protects Cyclin B from Degradation by Cdc20-Activated APC/C. Dev Cell 2019; 51:313-325.e10. [PMID: 31588029 DOI: 10.1016/j.devcel.2019.09.005] [Citation(s) in RCA: 52] [Impact Index Per Article: 8.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2019] [Revised: 06/26/2019] [Accepted: 09/05/2019] [Indexed: 12/23/2022]
Abstract
In the eukaryotic cell cycle, a threshold level of cyclin B accumulation triggers the G2-to-M transition, and subsequent cyclin B destruction triggers mitotic exit. The anaphase-promoting complex/cyclosome (APC/C) is the E3 ubiquitin ligase that, together with its co-activator Cdc20, targets cyclin B for destruction during mitotic exit. Here, we show that two pathways act in concert to protect cyclin B from Cdc20-activated APC/C in G2, in order to enable cyclin B accumulation and the G2-to-M transition. The first pathway involves the Mad1-Mad2 spindle checkpoint complex, acting in a distinct manner from checkpoint signaling after mitotic entry but employing a common molecular mechanism-the promotion of Mad2-Cdc20 complex formation. The second pathway involves cyclin-dependent kinase phosphorylation of Cdc20, which is known to reduce Cdc20's affinity for the APC/C. Cooperation of these two mechanisms, which target distinct APC/C binding interfaces of Cdc20, enables cyclin B accumulation and the G2-to-M transition.
Collapse
Affiliation(s)
- Pablo Lara-Gonzalez
- Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA, USA; Department of Cellular & Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA.
| | - Mark W Moyle
- Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA, USA; Department of Cellular & Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA
| | - Jacqueline Budrewicz
- Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA, USA; Department of Cellular & Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA
| | - Jose Mendoza-Lopez
- Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA, USA; Department of Cellular & Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA
| | - Karen Oegema
- Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA, USA; Department of Cellular & Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA
| | - Arshad Desai
- Ludwig Institute for Cancer Research, San Diego Branch, La Jolla, CA, USA; Department of Cellular & Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA.
| |
Collapse
|
|
38
|
Heestand B, Simon M, Frenk S, Titov D, Ahmed S. Transgenerational Sterility of Piwi Mutants Represents a Dynamic Form of Adult Reproductive Diapause. Cell Rep 2019; 23:156-171. [PMID: 29617657 PMCID: PMC5918633 DOI: 10.1016/j.celrep.2018.03.015] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2017] [Revised: 01/24/2018] [Accepted: 03/05/2018] [Indexed: 01/17/2023] Open
Abstract
Environmental stress can induce adult reproductive diapause, a state of developmental arrest that temporarily suspends reproduction. Deficiency for C. elegans Piwi protein PRG-1 results in strains that reproduce for many generations but then become sterile. We found that sterile-generation prg-1/Piwi mutants typically displayed pronounced germ cell atrophy as L4 larvae matured into 1-day-old adults. Atrophied germlines spontaneously reproliferated across the first days of adulthood, and this was accompanied by fertility for day 2–4 adults. Sterile day 5 prg-1 mutant adults remained sterile indefinitely, but providing an alternative food source could restore their fertility. Our data imply that late-generation prg-1 mutants experience a dynamic form of adult reproductive diapause, promoted by stress response, cell death, and RNAi pathways, where delayed fertility and reproductive quiescence represent parallel adaptive developmental outcomes. This may occur in response to a form of “heritable stress” that is transmitted by gametes and epigenetic in nature.
Collapse
Affiliation(s)
- Bree Heestand
- Department of Genetics, University of North Carolina, Chapel Hill, NC 27599, USA; Department of Biology, University of North Carolina, Chapel Hill, NC 27599, USA; Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599, USA
| | - Matt Simon
- Department of Genetics, University of North Carolina, Chapel Hill, NC 27599, USA; Department of Biology, University of North Carolina, Chapel Hill, NC 27599, USA; Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599, USA
| | - Stephen Frenk
- Department of Genetics, University of North Carolina, Chapel Hill, NC 27599, USA; Department of Biology, University of North Carolina, Chapel Hill, NC 27599, USA
| | - Denis Titov
- Department of Genetics, University of North Carolina, Chapel Hill, NC 27599, USA; Department of Biology, University of North Carolina, Chapel Hill, NC 27599, USA; Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599, USA
| | - Shawn Ahmed
- Department of Genetics, University of North Carolina, Chapel Hill, NC 27599, USA; Department of Biology, University of North Carolina, Chapel Hill, NC 27599, USA; Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599, USA; Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599, USA.
| |
Collapse
|
|
39
|
Drummond-Barbosa D. Local and Physiological Control of Germline Stem Cell Lineages in Drosophila melanogaster. Genetics 2019; 213:9-26. [PMID: 31488592 PMCID: PMC6727809 DOI: 10.1534/genetics.119.300234] [Citation(s) in RCA: 49] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/26/2019] [Accepted: 07/09/2019] [Indexed: 12/12/2022] Open
Abstract
The long-term survival of any multicellular species depends on the success of its germline in producing high-quality gametes and maximizing survival of the offspring. Studies in Drosophila melanogaster have led our growing understanding of how germline stem cell (GSC) lineages maintain their function and adjust their behavior according to varying environmental and/or physiological conditions. This review compares and contrasts the local regulation of GSCs by their specialized microenvironments, or niches; discusses how diet and diet-dependent factors, mating, and microorganisms modulate GSCs and their developing progeny; and briefly describes the tie between physiology and development during the larval phase of the germline cycle. Finally, it concludes with broad comparisons with other organisms and some future directions for further investigation.
Collapse
Affiliation(s)
- Daniela Drummond-Barbosa
- Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, Maryland 21205
| |
Collapse
|
|
40
|
Carranza-García E, Navarro RE. Apoptosis contributes to protect germ cells from the oogenic germline starvation response but is not essential for the gonad shrinking or recovery observed during adult reproductive diapause in C. elegans. PLoS One 2019; 14:e0218265. [PMID: 31194813 PMCID: PMC6564024 DOI: 10.1371/journal.pone.0218265] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2018] [Accepted: 05/29/2019] [Indexed: 12/18/2022] Open
Abstract
When C. elegans hermaphrodites are deprived of food during the mid-L4 larval stage and throughout adulthood, they enter an alternative stage termed "adult reproductive diapause (ARD)" in which they halt reproduction and extend their lifespan. During ARD, germ cell proliferation stops; oogenesis is slowed; and the gonad shrinks progressively, which has been described as the "oogenic germline starvation response". Upon refeeding, the shrunken gonad is regenerated, and animals recover fertility and live out their remaining lifespan. Little is known about the effects of ARD on oocyte quality after ARD. Thus, the aim of this study was to determine how oocyte quality is affected after ARD by measuring brood size and embryonic lethality as a reflection of defective oocyte production. We found that ARD affects reproductive capacity. The oogenic germline starvation response protects oogenic germ cells by slowing oogenesis to prevent prolonged arrest in diakinesis. In contrast to a previous report, we found that germ cell apoptosis is not the cause of gonad shrinkage; instead, we propose that ovulation contributes to gonad shrinkage during the oogenic germline starvation response. We show that germ cell apoptosis increases and continues during ARD via lin-35/Rb and an unknown mechanism. Although apoptosis contributes to maintain germ cell quality during ARD, we demonstrated that apoptosis is not essential to preserve animal fertility. Finally, we show that IIS signaling inactivation partially participates in the oogenic germline starvation response.
Collapse
Affiliation(s)
- E. Carranza-García
- Departamento de Biología Celular y Desarrollo, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Ciudad de México, México
| | - R. E. Navarro
- Departamento de Biología Celular y Desarrollo, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Ciudad de México, México
- * E-mail:
| |
Collapse
|
|
41
|
Cell Adhesion-Mediated Actomyosin Assembly Regulates the Activity of Cubitus Interruptus for Hematopoietic Progenitor Maintenance in Drosophila. Genetics 2019; 212:1279-1300. [PMID: 31138608 PMCID: PMC6707476 DOI: 10.1534/genetics.119.302209] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2018] [Accepted: 05/20/2019] [Indexed: 12/13/2022] Open
Abstract
The actomyosin network is involved in crucial cellular processes including morphogenesis, cell adhesion, apoptosis, proliferation, differentiation, and collective cell migration in Drosophila, Caenorhabditiselegans, and mammals. Here, we demonstrate that Drosophila larval blood stem-like progenitors require actomyosin activity for their maintenance. Genetic loss of the actomyosin network from progenitors caused a decline in their number. Likewise, the progenitor population increased upon sustained actomyosin activation via phosphorylation by Rho-associated kinase. We show that actomyosin positively regulates larval blood progenitors by controlling the maintenance factor Cubitus interruptus (Ci). Overexpression of the maintenance signal via a constitutively activated construct (ci.HA) failed to sustain Ci-155 in the absence of actomyosin components like Zipper (zip) and Squash (sqh), thus favoring protein kinase A (PKA)-independent regulation of Ci activity. Furthermore, we demonstrate that a change in cortical actomyosin assembly mediated by DE-cadherin modulates Ci activity, thereby determining progenitor status. Thus, loss of cell adhesion and downstream actomyosin activity results in desensitization of the progenitors to Hh signaling, leading to their differentiation. Our data reveal how cell adhesion and the actomyosin network cooperate to influence patterning, morphogenesis, and maintenance of the hematopoietic stem-like progenitor pool in the developing Drosophila hematopoietic organ.
Collapse
|
|
42
|
Mohammad K, Dakik P, Medkour Y, Mitrofanova D, Titorenko VI. Quiescence Entry, Maintenance, and Exit in Adult Stem Cells. Int J Mol Sci 2019; 20:ijms20092158. [PMID: 31052375 PMCID: PMC6539837 DOI: 10.3390/ijms20092158] [Citation(s) in RCA: 66] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2019] [Revised: 04/24/2019] [Accepted: 04/28/2019] [Indexed: 12/13/2022] Open
Abstract
Cells of unicellular and multicellular eukaryotes can respond to certain environmental cues by arresting the cell cycle and entering a reversible state of quiescence. Quiescent cells do not divide, but can re-enter the cell cycle and resume proliferation if exposed to some signals from the environment. Quiescent cells in mammals and humans include adult stem cells. These cells exhibit improved stress resistance and enhanced survival ability. In response to certain extrinsic signals, adult stem cells can self-renew by dividing asymmetrically. Such asymmetric divisions not only allow the maintenance of a population of quiescent cells, but also yield daughter progenitor cells. A multistep process of the controlled proliferation of these progenitor cells leads to the formation of one or more types of fully differentiated cells. An age-related decline in the ability of adult stem cells to balance quiescence maintenance and regulated proliferation has been implicated in many aging-associated diseases. In this review, we describe many traits shared by different types of quiescent adult stem cells. We discuss how these traits contribute to the quiescence, self-renewal, and proliferation of adult stem cells. We examine the cell-intrinsic mechanisms that allow establishing and sustaining the characteristic traits of adult stem cells, thereby regulating quiescence entry, maintenance, and exit.
Collapse
Affiliation(s)
- Karamat Mohammad
- Department of Biology, Concordia University, 7141 Sherbrooke Street, West, SP Building, Room 501-13, Montreal, QC H4B 1R6, Canada.
| | - Paméla Dakik
- Department of Biology, Concordia University, 7141 Sherbrooke Street, West, SP Building, Room 501-13, Montreal, QC H4B 1R6, Canada.
| | - Younes Medkour
- Department of Biology, Concordia University, 7141 Sherbrooke Street, West, SP Building, Room 501-13, Montreal, QC H4B 1R6, Canada.
| | - Darya Mitrofanova
- Department of Biology, Concordia University, 7141 Sherbrooke Street, West, SP Building, Room 501-13, Montreal, QC H4B 1R6, Canada.
| | - Vladimir I Titorenko
- Department of Biology, Concordia University, 7141 Sherbrooke Street, West, SP Building, Room 501-13, Montreal, QC H4B 1R6, Canada.
| |
Collapse
|
|
43
|
Kocsisova Z, Kornfeld K, Schedl T. Rapid population-wide declines in stem cell number and activity during reproductive aging in C. elegans. Development 2019; 146:dev173195. [PMID: 30936182 PMCID: PMC6503983 DOI: 10.1242/dev.173195] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2018] [Accepted: 03/13/2019] [Indexed: 01/03/2023]
Abstract
C. elegans hermaphrodites display dramatic age-related decline of reproduction early in life, while somatic functions are still robust. To understand reproductive aging, we analyzed the assembly line of oocyte production that generates fertilized eggs. Aging germlines displayed both sporadic and population-wide changes. A small fraction of aging animals displayed endomitotic oocytes in the germline and other defects. By contrast, all animals displayed age-related decreases in germline size and function. As early as day 3 of adulthood, animals displayed fewer stem cells and a slower cell cycle, which combine to substantially decrease progenitor zone output. The C. elegans germline is the only adult tissue that contains stem cells, allowing the analysis of stem cells in aging. To investigate the mechanism of the decrease in stem cell number, we analyzed the Notch signaling pathway. The Notch effectors LST-1 and SYGL-1 displayed age-related decreases in expression domains, suggesting a role for Notch signaling in germline aging. The results indicate that although sporadic defects account for the sterility of some animals, population-wide changes account for the overall pattern of reproductive aging.
Collapse
Affiliation(s)
- Zuzana Kocsisova
- Department of Developmental Biology, Washington University School of Medicine, St Louis, MO 63110, USA
- Department of Genetics, Washington University School of Medicine, St Louis, MO 63110, USA
| | - Kerry Kornfeld
- Department of Developmental Biology, Washington University School of Medicine, St Louis, MO 63110, USA
| | - Tim Schedl
- Department of Genetics, Washington University School of Medicine, St Louis, MO 63110, USA
| |
Collapse
|
|
44
|
Developmental Control of the Cell Cycle: Insights from Caenorhabditis elegans. Genetics 2019; 211:797-829. [PMID: 30846544 PMCID: PMC6404260 DOI: 10.1534/genetics.118.301643] [Citation(s) in RCA: 33] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2018] [Accepted: 10/10/2018] [Indexed: 12/11/2022] Open
Abstract
During animal development, a single fertilized egg forms a complete organism with tens to trillions of cells that encompass a large variety of cell types. Cell cycle regulation is therefore at the center of development and needs to be carried out in close coordination with cell differentiation, migration, and death, as well as tissue formation, morphogenesis, and homeostasis. The timing and frequency of cell divisions are controlled by complex combinations of external and cell-intrinsic signals that vary throughout development. Insight into how such controls determine in vivo cell division patterns has come from studies in various genetic model systems. The nematode Caenorhabditis elegans has only about 1000 somatic cells and approximately twice as many germ cells in the adult hermaphrodite. Despite the relatively small number of cells, C. elegans has diverse tissues, including intestine, nerves, striated and smooth muscle, and skin. C. elegans is unique as a model organism for studies of the cell cycle because the somatic cell lineage is invariant. Somatic cells divide at set times during development to produce daughter cells that adopt reproducible developmental fates. Studies in C. elegans have allowed the identification of conserved cell cycle regulators and provided insights into how cell cycle regulation varies between tissues. In this review, we focus on the regulation of the cell cycle in the context of C. elegans development, with reference to other systems, with the goal of better understanding how cell cycle regulation is linked to animal development in general.
Collapse
|
|
45
|
Palmisano NJ, Meléndez A. Autophagy in C. elegans development. Dev Biol 2019; 447:103-125. [PMID: 29709599 PMCID: PMC6204124 DOI: 10.1016/j.ydbio.2018.04.009] [Citation(s) in RCA: 32] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2017] [Revised: 03/19/2018] [Accepted: 04/12/2018] [Indexed: 12/11/2022]
Abstract
Autophagy involves the sequestration of cytoplasmic contents in a double-membrane structure referred to as the autophagosome and the degradation of its contents upon delivery to lysosomes. Autophagy activity has a role in multiple biological processes during the development of the nematode Caenorhabditis elegans. Basal levels of autophagy are required to remove aggregate prone proteins, paternal mitochondria, and spermatid-specific membranous organelles. During larval development, autophagy is required for the remodeling that occurs during dauer development, and autophagy can selectively degrade components of the miRNA-induced silencing complex, and modulate miRNA-mediated silencing. Basal levels of autophagy are important in synapse formation and in the germ line, to promote the proliferation of proliferating stem cells. Autophagy activity is also required for the efficient removal of apoptotic cell corpses by promoting phagosome maturation. Finally, autophagy is also involved in lipid homeostasis and in the aging process. In this review, we first describe the molecular complexes involved in the process of autophagy, its regulation, and mechanisms for cargo recognition. In the second section, we discuss the developmental contexts where autophagy has been shown to be important. Studies in C. elegans provide valuable insights into the physiological relevance of this process during metazoan development.
Collapse
Affiliation(s)
- Nicholas J Palmisano
- Biology Department, Queens College, CUNY, Flushing, NY, USA; Biology Ph.D. Program, The Graduate Center of the City University of New York, NK, USA
| | - Alicia Meléndez
- Biology Department, Queens College, CUNY, Flushing, NY, USA; Biology Ph.D. Program, The Graduate Center of the City University of New York, NK, USA; Biochemistry Ph.D. Program, The Graduate Center of the City University of New York, NY, USA.
| |
Collapse
|
|
46
|
|
|
47
|
Kocsisova Z, Mohammad A, Kornfeld K, Schedl T. Cell Cycle Analysis in the C. elegans Germline with the Thymidine Analog EdU. J Vis Exp 2018. [PMID: 30394383 DOI: 10.3791/58339] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023] Open
Abstract
Cell cycle analysis in eukaryotes frequently utilizes chromosome morphology, expression and/or localization of gene products required for various phases of the cell cycle, or the incorporation of nucleoside analogs. During S-phase, DNA polymerases incorporate thymidine analogs such as EdU or BrdU into chromosomal DNA, marking the cells for analysis. For C. elegans, the nucleoside analog EdU is fed to the worms during regular culture and is compatible with immunofluorescent techniques. The germline of C. elegans is a powerful model system for the studies of signaling pathways, stem cells, meiosis, and cell cycle because it is transparent, genetically facile, and meiotic prophase and cellular differentiation/gametogenesis occur in a linear assembly-like fashion. These features make EdU a great tool to study dynamic aspects of mitotically cycling cells and germline development. This protocol describes how to successfully prepare EdU bacteria, feed them to wild-type C. elegans hermaphrodites, dissect the hermaphrodite gonad, stain for EdU incorporation into DNA, stain with antibodies to detect various cell cycle and developmental markers, image the gonad and analyze the results. The protocol describes the variations in the method and analysis for the measurement of S-phase index, M-phase index, G2 duration, cell cycle duration, rate of meiotic entry, and rate of meiotic prophase progression. This method can be adapted to study the cell cycle or cell history in other tissues, stages, genetic backgrounds, and physiological conditions.
Collapse
Affiliation(s)
- Zuzana Kocsisova
- Department of Developmental Biology, Washington University School of Medicine, St. Louis, Missouri; Department of Genetics, Washington University School of Medicine, St. Louis, Missouri
| | - Ariz Mohammad
- Department of Genetics, Washington University School of Medicine, St. Louis, Missouri
| | - Kerry Kornfeld
- Department of Developmental Biology, Washington University School of Medicine, St. Louis, Missouri
| | - Tim Schedl
- Department of Genetics, Washington University School of Medicine, St. Louis, Missouri;
| |
Collapse
|
|
48
|
Seidel HS, Smith TA, Evans JK, Stamper JQ, Mast TG, Kimble J. C. elegans germ cells divide and differentiate in a folded tissue. Dev Biol 2018; 442:173-187. [DOI: 10.1016/j.ydbio.2018.07.013] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2018] [Revised: 07/08/2018] [Accepted: 07/16/2018] [Indexed: 12/18/2022]
|
|
49
|
Mohammad A, Vanden Broek K, Wang C, Daryabeigi A, Jantsch V, Hansen D, Schedl T. Initiation of Meiotic Development Is Controlled by Three Post-transcriptional Pathways in Caenorhabditis elegans. Genetics 2018; 209:1197-1224. [PMID: 29941619 PMCID: PMC6063227 DOI: 10.1534/genetics.118.300985] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2018] [Accepted: 06/20/2018] [Indexed: 11/18/2022] Open
Abstract
A major event in germline development is the transition from stem/progenitor cells to entry into meiosis and gametogenesis. This transition requires downregulation of mitotic cell cycle activity and upregulation of processes associated with meiosis. We identify the Caenorhabditis elegans SCFPROM-1 E3 ubiquitin-ligase complex as functioning to downregulate mitotic cell cycle protein levels including cyclin E, WAPL-1, and KNL-2 at meiotic entry and, independently, promoting homologous chromosome pairing as a positive regulator of the CHK-2 kinase. SCFPROM-1 is thus a novel regulator of meiotic entry, coordinating downregulation of mitotic cell cycle proteins and promoting homolog pairing. We further show that SCFPROM-1 functions redundantly, in parallel to the previously described GLD-1 and GLD-2 meiotic entry pathways, downstream of and inhibited by GLP-1 Notch signaling, which specifies the stem cell fate. Accordingly, C. elegans employs three post-transcriptional pathways, SCFPROM-1-mediated protein degradation, GLD-1-mediated translational repression, and GLD-2-mediated translational activation, to control and coordinate the initiation of meiotic development.
Collapse
Affiliation(s)
- Ariz Mohammad
- Department of Genetics, School of Medicine, Washington University in St. Louis, Missouri 63110
| | - Kara Vanden Broek
- Department of Biological Sciences, University of Calgary, T2N 1N4, Canada
| | - Christopher Wang
- Department of Biological Sciences, University of Calgary, T2N 1N4, Canada
| | - Anahita Daryabeigi
- Department of Chromosome Biology, Max F. Perutz Laboratories, University of Vienna, Vienna Biocenter, 1030, Austria
| | - Verena Jantsch
- Department of Chromosome Biology, Max F. Perutz Laboratories, University of Vienna, Vienna Biocenter, 1030, Austria
| | - Dave Hansen
- Department of Biological Sciences, University of Calgary, T2N 1N4, Canada
| | - Tim Schedl
- Department of Genetics, School of Medicine, Washington University in St. Louis, Missouri 63110
| |
Collapse
|
|
50
|
Foray V, Pérez-Jiménez MM, Fattouh N, Landmann F. Wolbachia Control Stem Cell Behavior and Stimulate Germline Proliferation in Filarial Nematodes. Dev Cell 2018; 45:198-211.e3. [PMID: 29689195 DOI: 10.1016/j.devcel.2018.03.017] [Citation(s) in RCA: 36] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2017] [Revised: 02/20/2018] [Accepted: 03/23/2018] [Indexed: 01/04/2023]
Abstract
Although symbiotic interactions are ubiquitous in the living world, examples of developmental symbioses are still scarce. We show here the crucial role of Wolbachia in the oogenesis of filarial nematodes, a class of parasites of biomedical and veterinary relevance. We applied newly developed techniques to demonstrate the earliest requirements of Wolbachia in the parasite germline preceding the production of faulty embryos in Wolbachia-depleted nematodes. We show that Wolbachia stimulate germline proliferation in a cell-autonomous manner, and not through nucleotide supplementation as previously hypothesized. We also found Wolbachia to maintain the quiescence of a pool of germline stem cells to ensure a constant delivery of about 1,400 eggs per day for many years. The loss of quiescence upon Wolbachia depletion as well as the disorganization of the distal germline suggest that Wolbachia are required to execute the proper germline stem cell developmental program in order to produce viable eggs and embryos.
Collapse
Affiliation(s)
- Vincent Foray
- CRBM, University of Montpellier, CNRS, Montpellier, France
| | | | - Nour Fattouh
- CRBM, University of Montpellier, CNRS, Montpellier, France
| | | |
Collapse
|