Ser7 of RNAPII-CTD facilitates heterochromatin formation by linking ncRNA to RNAi

Abstract

Some long noncoding RNAs (ncRNAs) transcribed by RNA polymerase II (RNAPII) are retained on chromatin, where they regulate RNAi and chromatin structure. The molecular basis of this retention remains unknown. We show that in fission yeast serine 7 (Ser7) of the C-terminal domain (CTD) of RNAPII is required for efficient siRNA generation for RNAi-dependent heterochromatin formation. Surprisingly, Ser7 facilitates chromatin retention of nascent heterochromatic RNAs (hRNAs). Chromatin retention of hRNAs and siRNA generation requires both Ser7 and an RNA-binding activity of the chromodomain of Chp1, a subunit of the RNA-induced transcriptional silencing (RITS) complex. Furthermore, RITS associates with RNAPII in a Ser7-dependent manner. We propose that Ser7 promotes cotranscriptional chromatin retention of hRNA by recruiting the RNA-chromatin connector protein Chp1, which facilitates RNAi-dependent heterochromatin formation. Our findings reveal a function of the CTD code: linking ncRNA transcription to RNAi for heterochromatin formation.

Heterochromatin suppresses gross chromosomal rearrangements at centromeres by repressing Tfs1/TFIIS-dependent transcription

“…9a). rpb1-S7A slightly increased dg and dh RNAs in otherwise wild-type background, as expected 72 . However, in clr4∆ cells, rpb1-S7A reduced the levels of centromeric noncoding RNAs most prominently at imr3 where transcription levels are low.…”

“…To obtain the direct evidence that RNAPII is involved in centromeric GCRs, we created the rpb1-S7A strain that harbored ChL. Consistent with low levels of H3K9me2 72 (Supplementary Fig. 8a), rpb1-S7A slightly increased GCR rate in otherwise wild-type background (Fig.…”

“…Ser7 of the CTD is required for transcription of noncoding small nuclear RNAs in human cells 70 . Changing all the Ser7 to Ala, rpb1-S7A , reduces chromatin-bound RNAs and H3K9me2 levels at fission yeast centromeres 71,72 . To obtain the direct evidence that RNAPII is involved in centromeric GCRs, we created the rpb1-S7A strain that harbored ChL.…”

Sfh1, an essential component of the RSC chromatin remodeling complex, maintains genome integrity in fission yeast

“…Transcripts from cnt::ura4 showed threefold reduction, while that of imr::ura4 showed about 1.4‐fold increase. As the levels of transcription at centromere region in wild‐type cell are quite low (about 1/1,000 of act1 gene, Figure d) and loss of heterochromatin caused 50‐ to 100‐fold increase in transcripts of marker genes inserted in centromeric heterochromatin (Kajitani et al., ), the observed changes of the amount of transcripts appear to be marginal. We also analyzed the localization of Cnp1 and H3K9me2 by ChIP assay to assess the effects of Sfh1 on centromeric chromatin structures (Figure e).…”

Trimethylguanosine synthase 1 (Tgs1) is involved in Swi6/HP1‐independent siRNA production and establishment of heterochromatin in fission yeast

“…Because precipitation of both antisense dg and dg RNA was reduced in tgs1Δ cells, we concluded that Tgs1 specifically hypermethylates dg and antisense dg RNA in both wild-type and swi6Δ cells. Previously, by analyzing RNAs immunoprecipitated with anti-histone H3 antibody, we showed that hncRNAs were retained on heterochromatin (Kajitani et al, 2017;Nakama et al, 2012). Hence, we examined the contribution of the TMG cap to chromatin binding by hncRNAs.…”