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1.  Dynein-dependent processive chromosome motions promote homologous pairing in C. elegans meiosis 
The Journal of Cell Biology  2012;196(1):47-64.
High-resolution time-lapse imaging of meiosis in C. elegans reveals stage-specific, dynein-driven chromosome motion that accelerates homologue pairing and triggers synapsis.
Meiotic chromosome segregation requires homologue pairing, synapsis, and crossover recombination, which occur during meiotic prophase. Telomere-led chromosome motion has been observed or inferred to occur during this stage in diverse species, but its mechanism and function remain enigmatic. In Caenorhabditis elegans, special chromosome regions known as pairing centers (PCs), rather than telomeres, associate with the nuclear envelope (NE) and the microtubule cytoskeleton. In this paper, we investigate chromosome dynamics in living animals through high-resolution four-dimensional fluorescence imaging and quantitative motion analysis. We find that chromosome movement is constrained before meiosis. Upon prophase onset, constraints are relaxed, and PCs initiate saltatory, processive, dynein-dependent motions along the NE. These dramatic motions are dispensable for homologous pairing and continue until synapsis is completed. These observations are consistent with the idea that motions facilitate pairing by enhancing the search rate but that their primary function is to trigger synapsis. This quantitative analysis of chromosome dynamics in a living animal extends our understanding of the mechanisms governing faithful genome inheritance.
doi:10.1083/jcb.201106022
PMCID: PMC3255982  PMID: 22232701
2.  Functional studies of the BTB domain in the Drosophila GAGA and Mod(mdg4) proteins 
Nucleic Acids Research  2000;28(20):3864-3870.
The BTB/POZ (BTB) domain is an approximately 120 residue sequence that is conserved at the N-terminus of many proteins in both vertebrates and invertebrates. We found that the protein encoded by a lethal allele of the Drosophila modifier of mdg4 [mod(mdg4)] gene has two mutated residues in its BTB domain. The identities of the residues at the positions of these mutations are highly conserved in the BTB domain family of proteins, and when the corresponding mutations were engineered into the BTB domain-containing GAGA protein, the activity of GAGA as a transcription activator in a transient transfection assay was severely reduced. The functional equivalence of the BTB domains was established by showing that the BTB domain of the mod(mdg4) protein can effectively substitute for that of GAGA.
PMCID: PMC110799  PMID: 11024164
3.  Comparative analysis of metazoan chromatin organization 
Ho, Joshua W. K. | Jung, Youngsook L. | Liu, Tao | Alver, Burak H. | Lee, Soohyun | Ikegami, Kohta | Sohn, Kyung-Ah | Minoda, Aki | Tolstorukov, Michael Y. | Appert, Alex | Parker, Stephen C. J. | Gu, Tingting | Kundaje, Anshul | Riddle, Nicole C. | Bishop, Eric | Egelhofer, Thea A. | Hu, Sheng’en Shawn | Alekseyenko, Artyom A. | Rechtsteiner, Andreas | Asker, Dalal | Belsky, Jason A. | Bowman, Sarah K. | Chen, Q. Brent | Chen, Ron A-J | Day, Daniel S. | Dong, Yan | Dose, Andrea C. | Duan, Xikun | Epstein, Charles B. | Ercan, Sevinc | Feingold, Elise A. | Ferrari, Francesco | Garrigues, Jacob M. | Gehlenborg, Nils | Good, Peter J. | Haseley, Psalm | He, Daniel | Herrmann, Moritz | Hoffman, Michael M. | Jeffers, Tess E. | Kharchenko, Peter V. | Kolasinska-Zwierz, Paulina | Kotwaliwale, Chitra V. | Kumar, Nischay | Langley, Sasha A. | Larschan, Erica N. | Latorre, Isabel | Libbrecht, Maxwell W. | Lin, Xueqiu | Park, Richard | Pazin, Michael J. | Pham, Hoang N. | Plachetka, Annette | Qin, Bo | Schwartz, Yuri B. | Shoresh, Noam | Stempor, Przemyslaw | Vielle, Anne | Wang, Chengyang | Whittle, Christina M. | Xue, Huiling | Kingston, Robert E. | Kim, Ju Han | Bernstein, Bradley E. | Dernburg, Abby F. | Pirrotta, Vincenzo | Kuroda, Mitzi I. | Noble, William S. | Tullius, Thomas D. | Kellis, Manolis | MacAlpine, David M. | Strome, Susan | Elgin, Sarah C. R. | Liu, Xiaole Shirley | Lieb, Jason D. | Ahringer, Julie | Karpen, Gary H. | Park, Peter J.
Nature  2014;512(7515):449-452.
doi:10.1038/nature13415
PMCID: PMC4227084  PMID: 25164756
4.  X-chromosome silencing in the germline of C. elegans 
Development (Cambridge, England)  2002;129(2):479-492.
SUMMARY
Germline maintenance in the nematode C. elegans requires global repressive mechanisms that involve chromatin organization. During meiosis, the X chromosome in both sexes exhibits a striking reduction of histone modifications that correlate with transcriptional activation when compared with the genome as a whole. The histone modification spectrum on the X chromosome corresponds with a lack of transcriptional competence, as measured by reporter transgene arrays. The X chromosome in XO males is structurally analogous to the sex body in mammals, contains a histone modification associated with heterochromatin in other species and is inactivated throughout meiosis. The synapsed X chromosomes in hermaphrodites also appear to be silenced in early meiosis, but genes on the X chromosome are detectably expressed at later stages of oocyte meiosis. Silencing of the sex chromosome during early meiosis is a conserved feature throughout the nematode phylum, and is not limited to hermaphroditic species.
PMCID: PMC4066729  PMID: 11807039
C. elegans; Germline; Silencing; X-inactivation; Histone modifications; Gametogenesis
5.  Identification of Chromosome Sequence Motifs That Mediate Meiotic Pairing and Synapsis in C. elegans 
Nature cell biology  2009;11(8):934-942.
C. elegans chromosomes contain specialized regions called pairing centers (PCs) that mediate homologous pairing and synapsis during meiosis. Four related proteins, ZIM-1, -2, -3, and HIM-8, associate with these sites and are required for their essential functions. Here we show that short sequence elements enriched in the corresponding chromosome regions selectively recruit these proteins in vivo. In vitro analysis using SELEX indicates that each protein’s binding specificity arises from a combination of two zinc fingers and an adjacent domain. Insertion of a cluster of recruiting motifs into a chromosome lacking its endogenous PC is sufficient to restore homologous pairing, synapsis, crossover recombination, and segregation. These findings help to illuminate how chromosome sites mediate essential aspects of meiotic chromosome dynamics.
doi:10.1038/ncb1904
PMCID: PMC4001799  PMID: 19620970
6.  Pushing the (nuclear) envelope into meiosis 
Genome Biology  2013;14(3):110.
A recent study shows that a short isoform of a mammalian nuclear lamin is important for homologous chromosome interactions during meiotic prophase in mice.
doi:10.1186/gb-2013-14-3-110
PMCID: PMC3663112  PMID: 23651872
7.  Chromosome pairing and synapsis during C. elegans meiosis 
Current opinion in cell biology  2013;25(3):349-356.
Meiosis is the specialized cell division cycle that produces haploid gametes to enable sexual reproduction. Reduction of chromosome number by half requires elaborate chromosome dynamics that occur in meiotic prophase to establish physical linkages between each pair of homologous chromosomes. C. elegans has emerged as an excellent model organism for molecular studies of meiosis, enabling investigators to combine the power of molecular genetics, cytology, and live analysis. Here we focus on recent studies that have shed light on how chromosomes find and identify their homologous partners, and the structural changes that accompany and mediate these interactions.
doi:10.1016/j.ceb.2013.03.003
PMCID: PMC3694717  PMID: 23578368
9.  The C. elegans DSB-2 Protein Reveals a Regulatory Network that Controls Competence for Meiotic DSB Formation and Promotes Crossover Assurance 
PLoS Genetics  2013;9(8):e1003674.
For most organisms, chromosome segregation during meiosis relies on deliberate induction of DNA double-strand breaks (DSBs) and repair of a subset of these DSBs as inter-homolog crossovers (COs). However, timing and levels of DSB formation must be tightly controlled to avoid jeopardizing genome integrity. Here we identify the DSB-2 protein, which is required for efficient DSB formation during C. elegans meiosis but is dispensable for later steps of meiotic recombination. DSB-2 localizes to chromatin during the time of DSB formation, and its disappearance coincides with a decline in RAD-51 foci marking early recombination intermediates and precedes appearance of COSA-1 foci marking CO-designated sites. These and other data suggest that DSB-2 and its paralog DSB-1 promote competence for DSB formation. Further, immunofluorescence analyses of wild-type gonads and various meiotic mutants reveal that association of DSB-2 with chromatin is coordinated with multiple distinct aspects of the meiotic program, including the phosphorylation state of nuclear envelope protein SUN-1 and dependence on RAD-50 to load the RAD-51 recombinase at DSB sites. Moreover, association of DSB-2 with chromatin is prolonged in mutants impaired for either DSB formation or formation of downstream CO intermediates. These and other data suggest that association of DSB-2 with chromatin is an indicator of competence for DSB formation, and that cells respond to a deficit of CO-competent recombination intermediates by prolonging the DSB-competent state. In the context of this model, we propose that formation of sufficient CO-competent intermediates engages a negative feedback response that leads to cessation of DSB formation as part of a major coordinated transition in meiotic prophase progression. The proposed negative feedback regulation of DSB formation simultaneously (1) ensures that sufficient DSBs are made to guarantee CO formation and (2) prevents excessive DSB levels that could have deleterious effects.
Author Summary
Formation of haploid gametes during meiosis relies on deliberate induction of DNA double-strand breaks (DSBs), followed by repair of a subset of DSBs as crossovers between homologous chromosomes. Crossovers form the basis of connections that enable homologs to segregate toward opposite spindle poles at meiosis I, thereby reducing ploidy. Thus, germ cells must generate enough DSBs to guarantee a crossover for every chromosome pair while avoiding an excessive number of DSBs that might endanger their genomes. Here, we provide insight into how this crucial balance is achieved. We identify C. elegans DSB-2 as a key regulator of DSB formation, and we propose that its association with chromatin is an indicator of DSB competence. Disappearance of DSB-2 is part of a coordinated transition affecting multiple distinct aspects of the meiotic program, and failure to form crossover-eligible recombination intermediates elicits a delay in DSB-2 removal and other transition events. Our data are consistent with a model in which meiotic DSB formation is governed by a negative feedback network wherein cells detect the presence of downstream crossover intermediates and respond by shutting down DSB formation, thereby ensuring that sufficient DSBs are made to guarantee crossovers while simultaneously minimizing the threat to genomic integrity.
doi:10.1371/journal.pgen.1003674
PMCID: PMC3738457  PMID: 23950729
10.  Identification of DSB-1, a Protein Required for Initiation of Meiotic Recombination in Caenorhabditis elegans, Illuminates a Crossover Assurance Checkpoint 
PLoS Genetics  2013;9(8):e1003679.
Meiotic recombination, an essential aspect of sexual reproduction, is initiated by programmed DNA double-strand breaks (DSBs). DSBs are catalyzed by the widely-conserved Spo11 enzyme; however, the activity of Spo11 is regulated by additional factors that are poorly conserved through evolution. To expand our understanding of meiotic regulation, we have characterized a novel gene, dsb-1, that is specifically required for meiotic DSB formation in the nematode Caenorhabditis elegans. DSB-1 localizes to chromosomes during early meiotic prophase, coincident with the timing of DSB formation. DSB-1 also promotes normal protein levels and chromosome localization of DSB-2, a paralogous protein that plays a related role in initiating recombination. Mutations that disrupt crossover formation result in prolonged DSB-1 association with chromosomes, suggesting that nuclei may remain in a DSB-permissive state. Extended DSB-1 localization is seen even in mutants with defects in early recombination steps, including spo-11, suggesting that the absence of crossover precursors triggers the extension. Strikingly, failure to form a crossover precursor on a single chromosome pair is sufficient to extend the localization of DSB-1 on all chromosomes in the same nucleus. Based on these observations we propose a model for crossover assurance that acts through DSB-1 to maintain a DSB-permissive state until all chromosome pairs acquire crossover precursors. This work identifies a novel component of the DSB machinery in C. elegans, and sheds light on an important pathway that regulates DSB formation for crossover assurance.
Author Summary
For most eukaryotes, recombination between homologous chromosomes during meiosis is an essential aspect of sexual reproduction. Meiotic recombination is initiated by programmed double-strand breaks in DNA, which have the potential to induce mutations if not efficiently repaired. To better understand the mechanisms that govern the initiation of recombination and regulate the formation of double-strand breaks, we use the nematode Caenorhabditis elegans as a model system. Here we describe a new gene, dsb-1, that is required for double-strand break formation in C. elegans. Through analysis of the encoded DSB-1 protein we illuminate an important regulatory pathway that promotes crossover recombination events on all chromosome pairs to ensure successful meiosis.
doi:10.1371/journal.pgen.1003679
PMCID: PMC3749324  PMID: 23990794
12.  Integrative Analysis of the Caenorhabditis elegans Genome by the modENCODE Project 
Gerstein, Mark B. | Lu, Zhi John | Van Nostrand, Eric L. | Cheng, Chao | Arshinoff, Bradley I. | Liu, Tao | Yip, Kevin Y. | Robilotto, Rebecca | Rechtsteiner, Andreas | Ikegami, Kohta | Alves, Pedro | Chateigner, Aurelien | Perry, Marc | Morris, Mitzi | Auerbach, Raymond K. | Feng, Xin | Leng, Jing | Vielle, Anne | Niu, Wei | Rhrissorrakrai, Kahn | Agarwal, Ashish | Alexander, Roger P. | Barber, Galt | Brdlik, Cathleen M. | Brennan, Jennifer | Brouillet, Jeremy Jean | Carr, Adrian | Cheung, Ming-Sin | Clawson, Hiram | Contrino, Sergio | Dannenberg, Luke O. | Dernburg, Abby F. | Desai, Arshad | Dick, Lindsay | Dosé, Andréa C. | Du, Jiang | Egelhofer, Thea | Ercan, Sevinc | Euskirchen, Ghia | Ewing, Brent | Feingold, Elise A. | Gassmann, Reto | Good, Peter J. | Green, Phil | Gullier, Francois | Gutwein, Michelle | Guyer, Mark S. | Habegger, Lukas | Han, Ting | Henikoff, Jorja G. | Henz, Stefan R. | Hinrichs, Angie | Holster, Heather | Hyman, Tony | Iniguez, A. Leo | Janette, Judith | Jensen, Morten | Kato, Masaomi | Kent, W. James | Kephart, Ellen | Khivansara, Vishal | Khurana, Ekta | Kim, John K. | Kolasinska-Zwierz, Paulina | Lai, Eric C. | Latorre, Isabel | Leahey, Amber | Lewis, Suzanna | Lloyd, Paul | Lochovsky, Lucas | Lowdon, Rebecca F. | Lubling, Yaniv | Lyne, Rachel | MacCoss, Michael | Mackowiak, Sebastian D. | Mangone, Marco | McKay, Sheldon | Mecenas, Desirea | Merrihew, Gennifer | Miller, David M. | Muroyama, Andrew | Murray, John I. | Ooi, Siew-Loon | Pham, Hoang | Phippen, Taryn | Preston, Elicia A. | Rajewsky, Nikolaus | Rätsch, Gunnar | Rosenbaum, Heidi | Rozowsky, Joel | Rutherford, Kim | Ruzanov, Peter | Sarov, Mihail | Sasidharan, Rajkumar | Sboner, Andrea | Scheid, Paul | Segal, Eran | Shin, Hyunjin | Shou, Chong | Slack, Frank J. | Slightam, Cindie | Smith, Richard | Spencer, William C. | Stinson, E. O. | Taing, Scott | Takasaki, Teruaki | Vafeados, Dionne | Voronina, Ksenia | Wang, Guilin | Washington, Nicole L. | Whittle, Christina M. | Wu, Beijing | Yan, Koon-Kiu | Zeller, Georg | Zha, Zheng | Zhong, Mei | Zhou, Xingliang | Ahringer, Julie | Strome, Susan | Gunsalus, Kristin C. | Micklem, Gos | Liu, X. Shirley | Reinke, Valerie | Kim, Stuart K. | Hillier, LaDeana W. | Henikoff, Steven | Piano, Fabio | Snyder, Michael | Stein, Lincoln | Lieb, Jason D. | Waterston, Robert H.
Science (New York, N.Y.)  2010;330(6012):1775-1787.
We systematically generated large-scale data sets to improve genome annotation for the nematode Caenorhabditis elegans, a key model organism. These data sets include transcriptome profiling across a developmental time course, genome-wide identification of transcription factor–binding sites, and maps of chromatin organization. From this, we created more complete and accurate gene models, including alternative splice forms and candidate noncoding RNAs. We constructed hierarchical networks of transcription factor–binding and microRNA interactions and discovered chromosomal locations bound by an unusually large number of transcription factors. Different patterns of chromatin composition and histone modification were revealed between chromosome arms and centers, with similarly prominent differences between autosomes and the X chromosome. Integrating data types, we built statistical models relating chromatin, transcription factor binding, and gene expression. Overall, our analyses ascribed putative functions to most of the conserved genome.
doi:10.1126/science.1196914
PMCID: PMC3142569  PMID: 21177976
13.  Cytoskeletal forces span the nuclear envelope to coordinate meiotic chromosome pairing and synapsis 
Cell  2009;139(5):907-919.
Summary
Sexual reproduction requires the unique cell division called meiosis, in which a diploid cell undergoes a reductional division to generate haploid gametes. A hallmark of meiotic prophase is the formation of pairwise linkages between homologous chromosomes, which later enable them to segregate from each other. In most organisms the pairing of homologous chromosomes is reinforced by synapsis, the polymerization of the synaptonemal complex (SC) between paired chromosome axes. The primary questions addressed here are: 1) how pairing is accomplished and 2) how synapsis is regulated so that it occurs selectively between homologs. We provide evidence that a connection between the chromosomes and the microtubule cytoskeleton via a bridge across the nuclear envelope is critical for both of these mechanisms. Our results indicate the existence of a mechanism that uses dynein to assess homology before licensing SC polymerization. The molecular components of this mechanism are conserved from fungi to mammals.
doi:10.1016/j.cell.2009.10.039
PMCID: PMC2825574  PMID: 19913287
14.  Correction: ZHP-3 Acts at Crossovers to Couple Meiotic Recombination with Synaptonemal Complex Disassembly and Bivalent Formation in C. elegans 
PLoS Genetics  2008;4(11):10.1371/annotation/ffbb52bd-7ceb-404c-8c96-93577bf83932.
doi:10.1371/annotation/ffbb52bd-7ceb-404c-8c96-93577bf83932
PMCID: PMC2598017
15.  ZHP-3 Acts at Crossovers to Couple Meiotic Recombination with Synaptonemal Complex Disassembly and Bivalent Formation in C. elegans 
PLoS Genetics  2008;4(10):e1000235.
Crossover recombination and the formation of chiasmata normally ensure the proper segregation of homologous chromosomes during the first meiotic division. zhp-3, the Caenorhabditis elegans ortholog of the budding yeast ZIP3 gene, is required for crossover recombination. We show that ZHP-3 protein localization is highly dynamic. At a key transition point in meiotic prophase, the protein shifts from along the length of the synaptonemal complex (SC) to an asymmetric localization on the SC and eventually becomes restricted to foci that mark crossover recombination events. A zhp-3::gfp transgene partially complements a null mutation and reveals a separation of function; although the fusion protein can promote nearly wild-type levels of recombination, aneuploidy among the progeny is high, indicating defects in meiotic chromosome segregation. The structure of bivalents is perturbed in this mutant, suggesting that the chromosome segregation defect results from an inability to properly remodel chromosomes in response to crossovers. smo-1 mutants exhibit phenotypes similar to zhp-3::gfp mutants at higher temperatures, and smo-1; zhp-3::gfp double mutants exhibit more severe meiotic defects than either single mutant, consistent with a role for SUMO in the process of SC disassembly and bivalent differentiation. We propose that coordination of crossover recombination with SC disassembly and bivalent formation reflects a conserved role of Zip3/ZHP-3 in coupling recombination with SC morphogenesis.
Author Summary
Sexual reproduction relies on meiosis. This specialized cell division generates gametes, such as sperm and eggs, with a single copy of the genome, so that fertilization restores diploidy. In order for chromosomes to segregate correctly during meiosis, homologs usually must undergo crossing over (genetic exchange) during meiotic prophase. How crossovers are coupled to large-scale changes in chromosome structure is not well understood. Our work shows that the protein ZHP-3 localizes to crossovers late in prophase, coincident with a transition in which chromosomes initiate progressive restructuring around the crossover. We have found that a ZHP-3-GFP fusion protein is competent to promote genetic exchange but not proper segregation. Chromosomes from these mutant animals exhibit defects in this late-prophase restructuring, suggesting that alterations in chromosome architecture that typically accompany crossovers have not occurred. We propose that ZHP-3 acts at crossovers to coordinate genetic exchange with higher order changes in chromosome structure that promote proper chromosome segregation.
doi:10.1371/journal.pgen.1000235
PMCID: PMC2567099  PMID: 18949042
16.  A Link between Meiotic Prophase Progression and Crossover Control 
PLoS Genetics  2006;2(2):e12.
During meiosis, most organisms ensure that homologous chromosomes undergo at least one exchange of DNA, or crossover, to link chromosomes together and accomplish proper segregation. How each chromosome receives a minimum of one crossover is unknown. During early meiosis in Caenorhabditis elegans and many other species, chromosomes adopt a polarized organization within the nucleus, which normally disappears upon completion of homolog synapsis. Mutations that impair synapsis even between a single pair of chromosomes in C. elegans delay this nuclear reorganization. We quantified this delay by developing a classification scheme for discrete stages of meiosis. Immunofluorescence localization of RAD-51 protein revealed that delayed meiotic cells also contained persistent recombination intermediates. Through genetic analysis, we found that this cytological delay in meiotic progression requires double-strand breaks and the function of the crossover-promoting heteroduplex HIM-14 (Msh4) and MSH-5. Failure of X chromosome synapsis also resulted in impaired crossover control on autosomes, which may result from greater numbers and persistence of recombination intermediates in the delayed nuclei. We conclude that maturation of recombination events on chromosomes promotes meiotic progression, and is coupled to the regulation of crossover number and placement. Our results have broad implications for the interpretation of meiotic mutants, as we have shown that asynapsis of a single chromosome pair can exert global effects on meiotic progression and recombination frequency.
Synopsis
Meiosis is a specialized cell division and an essential component of sexual reproduction. During meiotic prophase, each chromosome must pair with its unique homologous partner and undergo crossing over (genetic exchange) to segregate properly. A major mystery is how the molecular events of meiotic recombination are coupled to the large-scale dynamics of chromosome synapsis. This work reveals a link between the large-scale regulation of chromosome organization and the distribution of crossover events on the chromosomes. In C. elegans, defects in chromosome pairing or synapsis result in an extension of a normally transient stage of meiotic prophase. This study finds that this extension is associated with dysregulation of crossovers, so that more than the usual number of crossovers occur, and their distribution is shifted along the chromosomes. These observations contribute to our understanding of crossover control, which normally ensures accurate transmission of genetic information from parent to progeny.
doi:10.1371/journal.pgen.0020012
PMCID: PMC1359072  PMID: 16462941
17.  Here, There, and Everywhere 
The Journal of Cell Biology  2001;153(6):f33-f38.
PMCID: PMC2192025  PMID: 11402076

Results 1-17 (17)