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Differential spatial and structural organization of the X chromosome underlies dosage compensation in C. elegans.

ABSTRACT: The essential adaptation of X-linked gene expression to the X chromosome copy number variation (called dosage compensation, DC) has been widely studied as a model of chromosome-wide gene regulation. In C. elegans, DC is achieved by two fold downregulation of gene expression from both X copies in hermaphrodites. The dosage compensation complex (DCC), a multiprotein complex structurally similar to mitotic condensins, restricts RNA polymerase progression. Higher order chromatin structures have therefore long been suggested to regulate X-linked gene expression. Here we show that in C. elegans males, the single X chromosome interacts broadly with nuclear pores, while in hermaphrodites the DCC impairs this interaction. Using microscopic measurements we find that the X chromosome is located at the nuclear periphery in males and internal in hermaphrodites, while compaction was higher for the X chromosome than for autosomes but surprisingly comparable between sexes. Mechanistically, we show that a single motif enriched on X, sufficient for DCC loading in hermaphrodites, autonomously targets an autosomal locus to the nuclear periphery specifically in males. Using DNA adenine methyltransferase identification (DamID), we demonstrate that the perinuclear interaction domains are nuclear pores. Dynamic polymer modeling shows that this discrete pore anchoring can explain the high compaction of the male X chromosome. Together, our results put forward a structural model of DC, by demonstrating that X-specific sequences mediate interactions with nuclear pores, thereby locating the X chromosome in active perinuclear domains, while the DCC physically moves X-linked genes away from these transcriptionally active neighborhoods.

ORGANISM(S): Caenorhabditis elegans

PROVIDER: GSE56270 | GEO | 2014/12/01

SECONDARY ACCESSION(S): PRJNA242811

REPOSITORIES: GEO

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Project description:In Caenorhabditis elegans, the dosage compensation complex (DCC) specifically binds to and represses transcription from both X chromosomes in hermaphrodites. The DCC is composed of an X-specific condensin complex that interacts with several proteins. During embryogenesis, DCC starts localizing to the X chromosomes around the 40-cell stage, and is followed by X-enrichment of H4K20me1 between 100-cell to comma stage. Here, we analyzed dosage compensation of the X chromosome between sexes, and the roles of dpy-27 (condensin subunit), dpy-21 (non-condensin DCC member), set-1 (H4K20 monomethylase) and set-4 (H4K20 di-/tri-methylase) in X chromosome repression using mRNA-seq and ChIP-seq analyses across several developmental time points. We found that the DCC starts repressing the X chromosomes by the 40-cell stage, but X-linked transcript levels remain significantly higher in hermaphrodites compared to males through the comma stage of embryogenesis. Dpy-27 and dpy-21 are required for X chromosome repression throughout development, but particularly in early embryos dpy-27 and dpy-21 mutations produced distinct expression changes, suggesting a DCC independent role for dpy-21. We previously hypothesized that the DCC increases H4K20me1 by reducing set-4 activity on the X chromosomes. Accordingly, in the set-4 mutant, H4K20me1 increased more from the autosomes compared to the X, equalizing H4K20me1 level between X and autosomes. H4K20me1 increase on the autosomes led to a slight repression, resulting in a relative effect of X derepression. H4K20me1 depletion in the set-1 mutant showed greater X derepression compared to equalization of H4K20me1 levels between X and autosomes in the set-4 mutant, indicating that H4K20me1 level is important, but X to autosomal balance of H4K20me1 contributes only slightly to X-repression. Thus H4K20me1 by itself is not a downstream effector of the DCC. In summary, X chromosome dosage compensation starts in early embryos as the DCC localizes to the X, and is strengthened in later embryogenesis by H4K20me1.

Project description:The essential process of dosage compensation equalizes X-chromosome gene expression between C. elegans XO males and XX hermaphrodites through a dosage compensation complex (DCC) that resembles condensin. The DCC binds to both X chromosomes of hermaphrodites to repress transcription by half. Here we show that post-translational modification by the SUMO conjugation pathway is essential for sex-specific assembly of the DCC onto X. Depletion of the SUMO peptide in vivo severely disrupts binding of particular DCC subunits and causes changes in X-linked gene expression similar to those caused by disrupting genes encoding DCC subunits. Three DCC subunits are themselves SUMOylated, and depletion of SUMO preferentially reduces their binding to X, suggesting that SUMOylation of DCC subunits is essential for robust association with X. DCC SUMOylation is triggered by the signal that initiates DCC assembly onto X. The initial step of assembly--binding of X-targeting factors to recruitment sites on X (rex sites)--is independent of SUMOylation, but robust binding of the complete complex requires SUMOylation. SUMOylated DCC subunits are enriched at rex sites, and SUMOylation enhances interactions between X-targeting factors and condensin subunits that facilitate DCC binding beyond the low level achieved without SUMOylation. DCC subunits also participate in condensin complexes essential for chromosome segregation, but their SUMOylation occurs only in the context of the DCC. Our results reinforce a newly emerging theme in which multiple proteins of a complex are SUMOylated in response to a specific stimulus, leading to accelerated complex formation and enhanced function. ChIP-chip experiments using antibodies against DPY-27, SDC-3, DPY-30, DPY-26, DPY-28 and FLAG-tagged SDC-2 in wild-type and smo-1 RNAi treated mixed embryos, with IGG controls. Also, sequential ChIP-chip experiments: (1) ChIP using FLAG antibodies to determine the genome-wide binding sites for SUMOylated proteins, (2) ChIP using FLAG antibodies followed by re-ChIP of eluted protein-chromatin complexes with DPY-27 antibodies to determine genome-wide binding sites for SUMOylated DPY-27 (referred to as DPY-27 re-ChIP experiments), (3) ChIP using FLAG antibodies followed by re-ChIP of eluted protein-chromatin with IGG antibodies to determine background binding (referred to as IGG re-ChIP experiments, and (4) ChIP using DPY-27 antibodies as a control to assess the efficiency of DPY-27 binding and detection in control vs. FLAG-tagged strains.

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Differential spatial and structural organization of the X chromosome underlies dosage compensation in C. elegans.

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