Sgf73, a subunit of SAGA complex, is required for the assembly of RITS complex in fission yeast

Abstract

RNA interference (RNAi) is a widespread gene-silencing mechanism and is required for heterochromatin assembly in a variety of organisms. The RNA-induced transcriptional silencing complex (RITS), composed of Ago1, Tas3 and Chp1, is a key component of RNAi machinery in fission yeast that connects short interference RNA (siRNA) and heterochromatin formation. However, the process by which RITS is assembled is not well understood. Here, we identified Sgf73, a subunit of the SAGA co-transcriptional complex, is required for pericentromeric heterochromatin silencing and the generation of siRNA. This novel role of Sgf73 is independent of enzymatic activities or structural integrity of SAGA. Instead, Sgf73 is physically associated with Ago1 and Chp1. The interactions among the subunits of the RITS, including those between Tas3 and Chp1, between Chp1 and Ago1, between Ago1 and Tas3, were all impaired by the deletion of sgf73+. Consistently, the recruitment of Ago1 and Chp1 to the pericentromeric region was abolished in sgf73Δ cells. Our study unveils a moonlighting function of a SAGA subunit. It suggests Sgf73 is a novel factor that promotes assembly of RITS and RNAi-mediated heterochromatin formation.

Set3 contributes to heterochromatin integrity by promoting transcription of subunits of Clr4-Rik1-Cul4 histone methyltransferase complex in fission yeast

“…As H3K9me stabilizes the interaction between RITS and chromatin 10 , decreased H3K9me at pericentromeric region in the set3 Δ mutant might affect the proper recruitment of RITS and following siRNA production. The weak phenotype of set3 Δ mutant is contrast with the dramatic defects observed in the deletions of subunit of CLRC, or deletions of other factors that directly participate in the heterochromatin assembly 37 38 39 . Therefore, Set3 is not an on/off regulator, but is more like a fine-tuning factor contributing to the tight regulation of heterochromatin integrity at centromeres, telomeres and silent mating-type locus.…”

“…Northern blot was performed as described previously 39 . Primers and oligos used are listed in Supplementary Table S3 .…”

The molecular chaperone Hsp90 regulates heterochromatin assembly through stabilizing multiple complexes in fission yeast

“…Occasionally, some critical factors required for RNAi-dependent heterochromatin assembly, have been revealed by genetic screens or hint-directed genetic analyses. For example, the splicing factors Cwf10, Prp10 and Cwf14 (Bayne et al, 2008;Kallgren et al, 2014), the spliceosomal U4 snRNA (Chinen et al, 2010), the Spt-Ada-Gcn5-acetyltransferase (SAGA) complex subunit Sgf73 (Deng et al, 2015) and the inner nuclear membrane protein complex Lem2-Nur1 (Banday et al, 2016;Barrales et al, 2016) have been identified by these approaches. In this study, we provided another example where genetics helped to identify a novel factor, Hsp90, that broadly regulates the stability or assembly of factors or complexes involved in heterochromatin formation in S. pombe.…”

The promiscuity of the SAGA complex subunits: Multifunctional or moonlighting proteins?

“…As described for other SAGA subunits, Sgf73 has an alternative role that does not involve SAGA. For instance, in the fission yeast S. pombe Sgf73 functions as a factor necessary for the formation and function of the RNA-induced transcriptional silencing complex (RITS) [126]. RITS is a key component of the RNA interference (RNAi) gene silencing mechanism by which heterochromatin is assembled in many organisms.…”