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Nuclear aconitase regulates heterochromatin formation by interacting with Chp1

ABSTRACT: Aconitase is a highly conserved metabolic enzyme for TCA cycle. In Schizosaccharomyces pombe, an aconitase (Aco2) is localized in both mitochondria and the nucleus, apart from an exclusively mitochondrial one (Aco1). We investigated the role of nuclear Aco2, and present evidences that it interacts with Chp1, a heterochromatin-associated HP1 protein, and acts as an anti-silencing factor. Nuclear depletion of Aco2 by deleting nuclear localization signal (aco2ΔNLS) suppressed the gene-silencing defects of RNAi mutants Δago1 and Δdcr1 at the centromere, where RNAi-mediated heterochromatin formation prevails. The aco2 mutation caused an increase in Chp1 binding to the centromeric repeats, with concomitant increase in the level of dimethylated histone H3K9, reflecting restoration of heterochromatin formation. Co-precipitation revealed that Aco2 interacted directly with Chp1 via the N-terminal region (1-235 aa), where the chromodomain (20-75 aa) that recognizes methylated H3K9 is located. Genome-wide analysis revealed that the aco2ΔNLS mutation caused elevation in the level of H3K9me2 not only at heterochromatic but also at euchromatic regions in the Δago1 mutant background. We also found that the single aco2ΔNLS mutation caused aberrant gene silencing at sub-telomeric and some euchromatic coding genes. These results demonstrate that Aco2 inhibits heterochromatin assembly over a wide range, by interfering with Chp1. Its contribution is prominent in the RNAi-independent heterochromatic region or in the absence of RNAi effector complex RITS that holds Chp1. Taken together, this work shows a clear example of a metabolic enzyme that can function in the nucleus as an epigenetic modulator of gene expression.

ORGANISM(S): Schizosaccharomyces pombe

PROVIDER: GSE101804 | GEO | 2020/03/25

REPOSITORIES: GEO

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Project description:Heterochromatic DNA domains play important roles in the regulation of gene expression and maintenance of genome stability by silencing repetitive DNA elements and transposons. In organisms ranging from fission yeast to mammals, heterochromatin assembly at DNA repeats involves the activity of small noncoding RNAs (sRNAs) associated with the RNA interference (RNAi) pathway. Typically, sRNAs, originating from long noncoding RNAs transcribed from DNA repeats, guide Argonaute-containing effector complexes to complementary nascent RNAs to initiate histone H3 lysine 9 di- and tri-methylation (H3K9me2 and H3K9me3, respectively) and heterochromatin formation. H3K9me is in turn required for recruitment of RNAi to chromatin and promotes sRNA generation. However, how heterochromatin formation, which silences transcription, can proceed by a co-transcriptional mechanism that also promotes sRNA generation remains paradoxical. Here, using Clr4, the fission yeast homolog of mammalian SUV39H H3K9 methyltransferases, we designed active site mutations, which allow H3K9me2 catalysis but block the transition to H3K9me3. We show that H3K9me2 defines a functionally distinct heterochromatin state that is sufficient for RNAi-dependent co-transcriptional gene silencing (CTGS). Unlike H3K9me3 domains, which are transcriptionally silent, H3K9me2 domains are transcriptionally active, contain modifications associated with euchromatic transcription, and couple RNAi-mediated transcript degradation to the establishment of H3K9me domains. The two H3K9me states recruit reader proteins with different efficiencies, explaining their different downstream silencing functions. Furthermore, transition from H3K9me2 to H3K9me3 is required for RNAi-independent epigenetic inheritance of H3K9me domains. Our findings demonstrate that H3K9me2 and H3K9me3 define functionally distinct heterochromatin states and uncover a mechanism for formation of transcriptionally permissive heterochromatin that is compatible with its broadly conserved role in RNAi-mediated genome defense.

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Nuclear aconitase regulates heterochromatin formation by interacting with Chp1

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