Limits...
NatB domain-containing CRA-1 antagonizes hydrolase ACER-1 linking acetyl-CoA metabolism to the initiation of recombination during C. elegans meiosis.

Gao J, Kim HM, Elia AE, Elledge SJ, Colaiácovo MP - PLoS Genet. (2015)

Bottom Line: Moreover, perturbations to global histone acetylation levels are accompanied by changes in the frequency of DSB formation in C. elegans.CRA-1 is in turn negatively regulated by XND-1, an AT-hook containing protein.We propose that this newly defined protein network links acetyl-CoA metabolism to meiotic DSB formation via modulation of global histone acetylation.

View Article: PubMed Central - PubMed

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

ABSTRACT
The formation of DNA double-strand breaks (DSBs) must take place during meiosis to ensure the formation of crossovers, which are required for accurate chromosome segregation, therefore avoiding aneuploidy. However, DSB formation must be tightly regulated to maintain genomic integrity. How this regulation operates in the context of different chromatin architectures and accessibility, and how it is linked to metabolic pathways, is not understood. We show here that global histone acetylation levels undergo changes throughout meiotic progression. Moreover, perturbations to global histone acetylation levels are accompanied by changes in the frequency of DSB formation in C. elegans. We provide evidence that the regulation of histone acetylation requires CRA-1, a NatB domain-containing protein homologous to human NAA25, which controls the levels of acetyl-Coenzyme A (acetyl-CoA) by antagonizing ACER-1, a previously unknown and conserved acetyl-CoA hydrolase. CRA-1 is in turn negatively regulated by XND-1, an AT-hook containing protein. We propose that this newly defined protein network links acetyl-CoA metabolism to meiotic DSB formation via modulation of global histone acetylation.

Show MeSH

Related in: MedlinePlus

Expression and localization of CRA-1::GFP in the C. elegans germline.(A) CRA-1::GFP expression pattern in the adult hermaphrodite germline. Gonads dissected from CRA-1::GFP transgenic hermaphrodite worms were co-stained with anti-GFP antibody (green) and DAPI (blue). White arrowheads indicate entrance into meiosis (beginning of the transition zone). Bar, 20 μm. (B) High-magnification images of representative nuclei from different stages in the germline are shown to indicate CRA-1::GFP expression levels. Bar, 5 μm. (C) CRA-1::GFP expression in pachytene nuclei of gonads from either control(RNAi) (empty vector) or cra-1(RNAi) CRA-1::GFP transgenic worms. Gonads were stained with anti-GFP antibody (green) and DAPI (blue). Bar, 5 μm. (D) Western blot analysis of CRA-1::GFP and tubulin in control(RNAi) and cra-1(RNAi) CRA-1::GFP transgenic worms. (E) CRA-1::GFP is enriched on the autosomes. Pachytene nuclei were co-immunostained for CRA-1::GFP (green) and H3K36me3 (red). DNA was stained with DAPI (blue). Open arrowheads point to the X chromosomes. Bar, 4 μm. (F) Co-immunostaining for CRA-1::GFP (green) and SYP-1 (red) in pachytene nuclei of CRA-1::GFP transgenic worms. DNA was stained with DAPI (blue). Bar, 3 μm.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4359108&req=5

pgen.1005029.g002: Expression and localization of CRA-1::GFP in the C. elegans germline.(A) CRA-1::GFP expression pattern in the adult hermaphrodite germline. Gonads dissected from CRA-1::GFP transgenic hermaphrodite worms were co-stained with anti-GFP antibody (green) and DAPI (blue). White arrowheads indicate entrance into meiosis (beginning of the transition zone). Bar, 20 μm. (B) High-magnification images of representative nuclei from different stages in the germline are shown to indicate CRA-1::GFP expression levels. Bar, 5 μm. (C) CRA-1::GFP expression in pachytene nuclei of gonads from either control(RNAi) (empty vector) or cra-1(RNAi) CRA-1::GFP transgenic worms. Gonads were stained with anti-GFP antibody (green) and DAPI (blue). Bar, 5 μm. (D) Western blot analysis of CRA-1::GFP and tubulin in control(RNAi) and cra-1(RNAi) CRA-1::GFP transgenic worms. (E) CRA-1::GFP is enriched on the autosomes. Pachytene nuclei were co-immunostained for CRA-1::GFP (green) and H3K36me3 (red). DNA was stained with DAPI (blue). Open arrowheads point to the X chromosomes. Bar, 4 μm. (F) Co-immunostaining for CRA-1::GFP (green) and SYP-1 (red) in pachytene nuclei of CRA-1::GFP transgenic worms. DNA was stained with DAPI (blue). Bar, 3 μm.

Mentions: To further examine the link between changes in global histone acetylation and CRA-1 function we generated a transgenic line expressing a functional GFP tagged CRA-1 driven by a cra-1 promoter (Figs. 2, S1, S2). CRA-1::GFP is observed localizing in somatic and embryonic cells as well as to meiotic germline nuclei (Figs. 2A-B, S2A and S2B), suggesting that the role of CRA-1 may not be limited to meiosis. This is consistent with the elevated levels of larval lethality (61%) observed in cra-1 mutants [30]. The specificity of the observed CRA-1::GFP signal was confirmed by anti-GFP immunostaining of dissected gonads from transgenic worms depleted of CRA-1 by RNAi (Fig. 2C), and by western blotting (Fig. 2D). Analysis of CRA-1 localization during embryonic cell cycle progression does not show an obvious change of CRA-1 signal from interphase to mitotic prophase, although a signal reduction was observed from prometaphase to anaphase (S2C Fig.). In the germline, CRA-1 signal is first detected in early prophase nuclei (leptotene/zygotene stages), during which chromosomes reorganize spatially acquiring a crescent-shaped appearance (Fig. 2A-B). CRA-1 signal increases as meiotic nuclei progress into the pachytene stage, where chromosomes are fully synapsed and crossover formation is completed. Therefore, the impact of a cra-1 mutation on histone acetylation along the gonad is consistent with the pattern of expression observed for CRA-1, supporting the idea that CRA-1 may contribute to this dynamic acetylation. Moreover, the CRA-1::GFP transgene can also restore histone acetylation in cra-1 mutants (S1E Fig.). Therefore, these data are consistent with a role for CRA-1 as a positive regulator of global histone acetylation.


NatB domain-containing CRA-1 antagonizes hydrolase ACER-1 linking acetyl-CoA metabolism to the initiation of recombination during C. elegans meiosis.

Gao J, Kim HM, Elia AE, Elledge SJ, Colaiácovo MP - PLoS Genet. (2015)

Expression and localization of CRA-1::GFP in the C. elegans germline.(A) CRA-1::GFP expression pattern in the adult hermaphrodite germline. Gonads dissected from CRA-1::GFP transgenic hermaphrodite worms were co-stained with anti-GFP antibody (green) and DAPI (blue). White arrowheads indicate entrance into meiosis (beginning of the transition zone). Bar, 20 μm. (B) High-magnification images of representative nuclei from different stages in the germline are shown to indicate CRA-1::GFP expression levels. Bar, 5 μm. (C) CRA-1::GFP expression in pachytene nuclei of gonads from either control(RNAi) (empty vector) or cra-1(RNAi) CRA-1::GFP transgenic worms. Gonads were stained with anti-GFP antibody (green) and DAPI (blue). Bar, 5 μm. (D) Western blot analysis of CRA-1::GFP and tubulin in control(RNAi) and cra-1(RNAi) CRA-1::GFP transgenic worms. (E) CRA-1::GFP is enriched on the autosomes. Pachytene nuclei were co-immunostained for CRA-1::GFP (green) and H3K36me3 (red). DNA was stained with DAPI (blue). Open arrowheads point to the X chromosomes. Bar, 4 μm. (F) Co-immunostaining for CRA-1::GFP (green) and SYP-1 (red) in pachytene nuclei of CRA-1::GFP transgenic worms. DNA was stained with DAPI (blue). Bar, 3 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4359108&req=5

pgen.1005029.g002: Expression and localization of CRA-1::GFP in the C. elegans germline.(A) CRA-1::GFP expression pattern in the adult hermaphrodite germline. Gonads dissected from CRA-1::GFP transgenic hermaphrodite worms were co-stained with anti-GFP antibody (green) and DAPI (blue). White arrowheads indicate entrance into meiosis (beginning of the transition zone). Bar, 20 μm. (B) High-magnification images of representative nuclei from different stages in the germline are shown to indicate CRA-1::GFP expression levels. Bar, 5 μm. (C) CRA-1::GFP expression in pachytene nuclei of gonads from either control(RNAi) (empty vector) or cra-1(RNAi) CRA-1::GFP transgenic worms. Gonads were stained with anti-GFP antibody (green) and DAPI (blue). Bar, 5 μm. (D) Western blot analysis of CRA-1::GFP and tubulin in control(RNAi) and cra-1(RNAi) CRA-1::GFP transgenic worms. (E) CRA-1::GFP is enriched on the autosomes. Pachytene nuclei were co-immunostained for CRA-1::GFP (green) and H3K36me3 (red). DNA was stained with DAPI (blue). Open arrowheads point to the X chromosomes. Bar, 4 μm. (F) Co-immunostaining for CRA-1::GFP (green) and SYP-1 (red) in pachytene nuclei of CRA-1::GFP transgenic worms. DNA was stained with DAPI (blue). Bar, 3 μm.
Mentions: To further examine the link between changes in global histone acetylation and CRA-1 function we generated a transgenic line expressing a functional GFP tagged CRA-1 driven by a cra-1 promoter (Figs. 2, S1, S2). CRA-1::GFP is observed localizing in somatic and embryonic cells as well as to meiotic germline nuclei (Figs. 2A-B, S2A and S2B), suggesting that the role of CRA-1 may not be limited to meiosis. This is consistent with the elevated levels of larval lethality (61%) observed in cra-1 mutants [30]. The specificity of the observed CRA-1::GFP signal was confirmed by anti-GFP immunostaining of dissected gonads from transgenic worms depleted of CRA-1 by RNAi (Fig. 2C), and by western blotting (Fig. 2D). Analysis of CRA-1 localization during embryonic cell cycle progression does not show an obvious change of CRA-1 signal from interphase to mitotic prophase, although a signal reduction was observed from prometaphase to anaphase (S2C Fig.). In the germline, CRA-1 signal is first detected in early prophase nuclei (leptotene/zygotene stages), during which chromosomes reorganize spatially acquiring a crescent-shaped appearance (Fig. 2A-B). CRA-1 signal increases as meiotic nuclei progress into the pachytene stage, where chromosomes are fully synapsed and crossover formation is completed. Therefore, the impact of a cra-1 mutation on histone acetylation along the gonad is consistent with the pattern of expression observed for CRA-1, supporting the idea that CRA-1 may contribute to this dynamic acetylation. Moreover, the CRA-1::GFP transgene can also restore histone acetylation in cra-1 mutants (S1E Fig.). Therefore, these data are consistent with a role for CRA-1 as a positive regulator of global histone acetylation.

Bottom Line: Moreover, perturbations to global histone acetylation levels are accompanied by changes in the frequency of DSB formation in C. elegans.CRA-1 is in turn negatively regulated by XND-1, an AT-hook containing protein.We propose that this newly defined protein network links acetyl-CoA metabolism to meiotic DSB formation via modulation of global histone acetylation.

View Article: PubMed Central - PubMed

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

ABSTRACT
The formation of DNA double-strand breaks (DSBs) must take place during meiosis to ensure the formation of crossovers, which are required for accurate chromosome segregation, therefore avoiding aneuploidy. However, DSB formation must be tightly regulated to maintain genomic integrity. How this regulation operates in the context of different chromatin architectures and accessibility, and how it is linked to metabolic pathways, is not understood. We show here that global histone acetylation levels undergo changes throughout meiotic progression. Moreover, perturbations to global histone acetylation levels are accompanied by changes in the frequency of DSB formation in C. elegans. We provide evidence that the regulation of histone acetylation requires CRA-1, a NatB domain-containing protein homologous to human NAA25, which controls the levels of acetyl-Coenzyme A (acetyl-CoA) by antagonizing ACER-1, a previously unknown and conserved acetyl-CoA hydrolase. CRA-1 is in turn negatively regulated by XND-1, an AT-hook containing protein. We propose that this newly defined protein network links acetyl-CoA metabolism to meiotic DSB formation via modulation of global histone acetylation.

Show MeSH
Related in: MedlinePlus