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A yeast two-hybrid screen for SYP-3 interactors identifies SYP-4, a component required for synaptonemal complex assembly and chiasma formation in Caenorhabditis elegans meiosis.

Smolikov S, Schild-Prüfert K, Colaiácovo MP - PLoS Genet. (2009)

Bottom Line: SYP-4 is essential for the localization of SYP-1, SYP-2, and SYP-3 CR proteins onto chromosomes, thereby playing a crucial role in the stabilization of pairing interactions between homologous chromosomes.The lack of chiasmata observed in syp-4 mutants supports the elevated levels of chromosome nondisjunction manifested in high embryonic lethality.Altogether our findings place SYP-4 as a central player in SC formation and broaden our understanding of the structure of the SC and its assembly.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
The proper assembly of the synaptonemal complex (SC) between homologs is critical to ensure accurate meiotic chromosome segregation. The SC is a meiotic tripartite structure present from yeast to humans, comprised of proteins assembled along the axes of the chromosomes and central region (CR) proteins that bridge the two chromosome axes. Here we identify SYP-4 as a novel structural component of the SC in Caenorhabditis elegans. SYP-4 interacts in a yeast two-hybrid assay with SYP-3, one of components of the CR of the SC, and is localized at the interface between homologs during meiosis. SYP-4 is essential for the localization of SYP-1, SYP-2, and SYP-3 CR proteins onto chromosomes, thereby playing a crucial role in the stabilization of pairing interactions between homologous chromosomes. In the absence of SYP-4, the levels of recombination intermediates, as indicated by RAD-51 foci, are elevated in mid-prophase nuclei, and crossover recombination events are significantly reduced. The lack of chiasmata observed in syp-4 mutants supports the elevated levels of chromosome nondisjunction manifested in high embryonic lethality. Altogether our findings place SYP-4 as a central player in SC formation and broaden our understanding of the structure of the SC and its assembly.

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Related in: MedlinePlus

Progression of meiotic recombination is impaired in syp-4 mutants.(A) Diagram of a C. elegans germline depicting the seven zones throughout which RAD-51 foci were scored for all nuclei. Levels of RAD-51 foci are indicated by the color code. (B) High magnification images of pachytene nuclei in wild type and syp-4 mutants stained with DAPI and RAD-51. Elevated levels of RAD-51 foci are observed on pachytene nuclei in syp-4 mutants. Bars, 2 µm. (C) Histograms depict the quantitation of RAD-51 foci in wild type and syp-4 mutant germlines. The percentage of nuclei observed for each category indicated by the color code (y-axis), are depicted for each zone along the germline (x-axis).
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pgen-1000669-g006: Progression of meiotic recombination is impaired in syp-4 mutants.(A) Diagram of a C. elegans germline depicting the seven zones throughout which RAD-51 foci were scored for all nuclei. Levels of RAD-51 foci are indicated by the color code. (B) High magnification images of pachytene nuclei in wild type and syp-4 mutants stained with DAPI and RAD-51. Elevated levels of RAD-51 foci are observed on pachytene nuclei in syp-4 mutants. Bars, 2 µm. (C) Histograms depict the quantitation of RAD-51 foci in wild type and syp-4 mutant germlines. The percentage of nuclei observed for each category indicated by the color code (y-axis), are depicted for each zone along the germline (x-axis).

Mentions: Interhomolog recombination resulting in crossover events is dependent on chromosome synapsis [2],[13],[14]. Therefore, we examined the progression of meiotic recombination in the syp-4(tm2713) mutants by immunostaining whole mounted germlines with an anti-RAD-51 antibody (RAD-51 is required for strand invasion/exchange during double-strand break repair; [39]). Specifically, wild type and syp-4(tm2713) mutant germlines were divided into 7 zones and levels of RAD-51 foci/nucleus were quantitated for all nuclei in each zone (Figure 6A). In wild type gonads, levels of RAD-51 foci started to increase as nuclei entered into meiotic prophase and peaked in early to mid pachytene (Figure 6C, zone 5, 3.7 foci/nucleus), after which they gradually declined (Figure 6C, zone 7, 0.5 foci/nucleus). As nuclei exited pachytene, RAD-51 foci were no longer observed. In syp-4(tm2713) mutants, the increase in the levels of RAD-51 foci was first observed with a similar timing to wild-type, suggesting that the initiation of meiotic recombination is not dependent on SYP-4 (Figure 6C). However, levels of RAD-51 foci were significantly higher in mid-pachytene in syp-4(tm2713) mutants compared to wild type (Figure 6B and 6C, zone 5, 10.2 foci/nucleus; p<0.0001, two-tailed Mann-Whitney test, 95% C.I.) and remained elevated up to late pachytene (Figure 6C, zone 7, 2.1 foci/nucleus; p<0.0001, two-tailed Mann-Whitney test, 95% C.I.). The defect in DSB repair progression observed in syp-4(tm2713) mutants probably stems from the lack of chromosome synapsis and therefore a lack of close and stable proximity to a homologous template for repair. However, DSB repair is eventually accomplished in syp-4(tm2713) mutants, as RAD-51 foci were absent in diplotene nuclei and chromosome fragments were not apparent in oocytes at diakinesis (Figure 2H). This delayed repair may proceed in part through recombination with the sister chromatid, as previously demonstrated for syp-2 and syp-3 mutants [13],[40].


A yeast two-hybrid screen for SYP-3 interactors identifies SYP-4, a component required for synaptonemal complex assembly and chiasma formation in Caenorhabditis elegans meiosis.

Smolikov S, Schild-Prüfert K, Colaiácovo MP - PLoS Genet. (2009)

Progression of meiotic recombination is impaired in syp-4 mutants.(A) Diagram of a C. elegans germline depicting the seven zones throughout which RAD-51 foci were scored for all nuclei. Levels of RAD-51 foci are indicated by the color code. (B) High magnification images of pachytene nuclei in wild type and syp-4 mutants stained with DAPI and RAD-51. Elevated levels of RAD-51 foci are observed on pachytene nuclei in syp-4 mutants. Bars, 2 µm. (C) Histograms depict the quantitation of RAD-51 foci in wild type and syp-4 mutant germlines. The percentage of nuclei observed for each category indicated by the color code (y-axis), are depicted for each zone along the germline (x-axis).
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Related In: Results  -  Collection

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pgen-1000669-g006: Progression of meiotic recombination is impaired in syp-4 mutants.(A) Diagram of a C. elegans germline depicting the seven zones throughout which RAD-51 foci were scored for all nuclei. Levels of RAD-51 foci are indicated by the color code. (B) High magnification images of pachytene nuclei in wild type and syp-4 mutants stained with DAPI and RAD-51. Elevated levels of RAD-51 foci are observed on pachytene nuclei in syp-4 mutants. Bars, 2 µm. (C) Histograms depict the quantitation of RAD-51 foci in wild type and syp-4 mutant germlines. The percentage of nuclei observed for each category indicated by the color code (y-axis), are depicted for each zone along the germline (x-axis).
Mentions: Interhomolog recombination resulting in crossover events is dependent on chromosome synapsis [2],[13],[14]. Therefore, we examined the progression of meiotic recombination in the syp-4(tm2713) mutants by immunostaining whole mounted germlines with an anti-RAD-51 antibody (RAD-51 is required for strand invasion/exchange during double-strand break repair; [39]). Specifically, wild type and syp-4(tm2713) mutant germlines were divided into 7 zones and levels of RAD-51 foci/nucleus were quantitated for all nuclei in each zone (Figure 6A). In wild type gonads, levels of RAD-51 foci started to increase as nuclei entered into meiotic prophase and peaked in early to mid pachytene (Figure 6C, zone 5, 3.7 foci/nucleus), after which they gradually declined (Figure 6C, zone 7, 0.5 foci/nucleus). As nuclei exited pachytene, RAD-51 foci were no longer observed. In syp-4(tm2713) mutants, the increase in the levels of RAD-51 foci was first observed with a similar timing to wild-type, suggesting that the initiation of meiotic recombination is not dependent on SYP-4 (Figure 6C). However, levels of RAD-51 foci were significantly higher in mid-pachytene in syp-4(tm2713) mutants compared to wild type (Figure 6B and 6C, zone 5, 10.2 foci/nucleus; p<0.0001, two-tailed Mann-Whitney test, 95% C.I.) and remained elevated up to late pachytene (Figure 6C, zone 7, 2.1 foci/nucleus; p<0.0001, two-tailed Mann-Whitney test, 95% C.I.). The defect in DSB repair progression observed in syp-4(tm2713) mutants probably stems from the lack of chromosome synapsis and therefore a lack of close and stable proximity to a homologous template for repair. However, DSB repair is eventually accomplished in syp-4(tm2713) mutants, as RAD-51 foci were absent in diplotene nuclei and chromosome fragments were not apparent in oocytes at diakinesis (Figure 2H). This delayed repair may proceed in part through recombination with the sister chromatid, as previously demonstrated for syp-2 and syp-3 mutants [13],[40].

Bottom Line: SYP-4 is essential for the localization of SYP-1, SYP-2, and SYP-3 CR proteins onto chromosomes, thereby playing a crucial role in the stabilization of pairing interactions between homologous chromosomes.The lack of chiasmata observed in syp-4 mutants supports the elevated levels of chromosome nondisjunction manifested in high embryonic lethality.Altogether our findings place SYP-4 as a central player in SC formation and broaden our understanding of the structure of the SC and its assembly.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
The proper assembly of the synaptonemal complex (SC) between homologs is critical to ensure accurate meiotic chromosome segregation. The SC is a meiotic tripartite structure present from yeast to humans, comprised of proteins assembled along the axes of the chromosomes and central region (CR) proteins that bridge the two chromosome axes. Here we identify SYP-4 as a novel structural component of the SC in Caenorhabditis elegans. SYP-4 interacts in a yeast two-hybrid assay with SYP-3, one of components of the CR of the SC, and is localized at the interface between homologs during meiosis. SYP-4 is essential for the localization of SYP-1, SYP-2, and SYP-3 CR proteins onto chromosomes, thereby playing a crucial role in the stabilization of pairing interactions between homologous chromosomes. In the absence of SYP-4, the levels of recombination intermediates, as indicated by RAD-51 foci, are elevated in mid-prophase nuclei, and crossover recombination events are significantly reduced. The lack of chiasmata observed in syp-4 mutants supports the elevated levels of chromosome nondisjunction manifested in high embryonic lethality. Altogether our findings place SYP-4 as a central player in SC formation and broaden our understanding of the structure of the SC and its assembly.

Show MeSH
Related in: MedlinePlus