Abstract
Heterochromatin formation in fission yeast depends on RNAi machinery and histone-modifying enzymes. One of the key histone-modifying complexes is Clr4-Rik1-Cul4 methyltransferase complex (CLRC), which mediates histone H3K9 methylation, a hallmark for heterochromatin. CLRC is composed of the Clr4 histone methyltransferase, Rik1, Raf1, Raf2 and Pcu4. However, transcriptional regulation of the CLRC subunits is not well understood. In this study, we identified Set3, a core subunit of the Set3/Hos2 histone deacetylase complex (Set3C), as a contributor to the integrity and silencing of heterochromatin at centromeres, telomeres and silent mating-type locus. This novel role of Set3 relies on its PHD finger, but is independent of deacetylase activity or structural integrity of Set3C. Set3 is not located to the centromeric region. Instead, Set3 is targeted to the promoters of clr4+ and rik1+, probably through its PHD finger. Set3 promotes transcription of clr4+ and rik1+. Consistently, the protein levels of Clr4 and Rik1 were reduced in the set3Δ mutant. The heterochromatin silencing defect in the set3Δ mutant could be rescued by overexpressing of clr4+ or rik1+. Our study suggests transcriptional activation of essential heterochromatin factors underlies the tight regulation of heterochromatin integrity.
Highlights
Cul4 methyltransferase complex (CLRC) consists of Clr[4], the β-propeller protein Rik[1], the cullin protein Pcu[4], the WD-40 protein Raf[1] and the Zn-finger protein Raf[2][14,15,16,17,18]
To dissect the mechanism of heterochromatin assembly, we carried out a genetic screen for mutants that displayed a defect in centromeric silencing in fission yeast
We used a parental strain in which the native ura4+ gene was deleted and a ura4+ marker gene was inserted into the outermost pericentromeric heterochromatin of chromosome 1 (Fig. 1a)[29]
Summary
CLRC consists of Clr[4], the β-propeller protein Rik[1], the cullin protein Pcu[4] ( known as Cul4), the WD-40 protein Raf[1] ( known as Dos1/Clr8/Cmc1) and the Zn-finger protein Raf[2] ( known as Dos2/Clr7/ Cmc2)[14,15,16,17,18]. Set[3] was first characterized in budding yeast by its feature of containing PHD finger and SET domain[21] The combination of both domains is a characteristic displayed by a group of trx-G proteins, including histone methyltransferase Trx and Ash[122]. We show that Set[3] contributes to the integrity of heterochromatin by promoting the transcription of Clr[4] and Rik[1], two key subunits of CLRC. This role of Set[3] is independent of Set3C, indicating a novel way of Set3-mediated transcriptional regulation
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