Contrasting roles of the RSC and ISW1/CHD1 chromatin remodelers in RNA polymerase II elongation and termination

Abstract

Most yeast genes have a nucleosome-depleted region (NDR) at the promoter and an array of regularly spaced nucleosomes phased relative to the transcription start site. We have examined the interplay between RSC (a conserved essential SWI/SNFtype complex that determines NDR size) and the ISW1, CHD1, and ISW2 nucleosome spacing enzymes in chromatin organization and transcription, using isogenic strains lacking all combinations of these enzymes. The contributions of these remodelers to chromatin organization are largely combinatorial, distinct, and nonredundant, supporting a model in which the +1 nucleosome is positioned by RSC and then used as a reference nucleosome by the spacing enzymes. Defective chromatin organization correlates with altered RNA polymerase II (Pol II) distribution. RSC-depleted cells exhibit low levels of elongating Pol II and high levels of terminating Pol II, consistent with defects in both termination and initiation, suggesting that RSC facilitates both. Cells lacking both ISW1 and CHD1 show the opposite Pol II distribution, suggesting elongation and termination defects. These cells have extremely disrupted chromatin, with high levels of closely packed dinucleosomes involving the second (+2) nucleosome. We propose that ISW1 and CHD1 facilitate Pol II elongation by separating closely packed nucleosomes.

The RSC complex remodels nucleosomes in transcribed coding sequences and promotes transcription inSaccharomyces cerevisiae

“…To test the above prediction, we examined the Pol II ChIP-seq data from isogenic WT and Rsc8depleted cells (rsc8), published by Clark and colleagues (Ocampo et al, 2019). We observed that the reductions in Pol II occupancies upon Rsc8 depletion were greatest at genes displaying the highest Pol II occupancies in WT cells, genome-wide ( Figure 4D and 4E).…”

“…Consistent with this, loss of RSC function resulted in NDR filling genome-wide in a manner independent of transcription status ( Figures 4C-4E). Immunoprecipitation-independent methods such as "CUT&RUN" (Skene and Henikoff, 2017) and ChEC-seq have shown that RSC localizes to -1 and +1 nucleosomes for the majority of genes, and that RSC loss leads to the movement of -1 and +1 nucleosomes towards NDRs genome-wide (Brahma and Henikoff, 2019;Klein-Brill et al, 2019;Kubik et al, 2019;Kubik et al, 2018;Ocampo et al, 2019;Parnell et al, 2015). These methods detected RSC signal at more genes than conventional ChIP, providing evidence that RSC may indeed interact transiently with promoter nucleosomes at these genes.…”

“…In contrast, rsc8 elicited a relatively small effect on the NDR filling for the highly transcribed Pol II-T10 genes ( Figure 4G), despite these genes showing the greatest reductions in Pol II occupancies in rsc8 cells. Depletion of Rsc8 was shown to be associated with increases in Pol II occupancies towards the 3'ends of transcribed genes (Ocampo et al, 2019) (Figure S4C and S4D). Although a relatively minor shift in +1 nucleosomes in rsc8 could lower Pol II occupancies in CDSs ( Figure 4F), but it does not fully explain the altered profile of Pol II observed in rsc8 cells.…”

“…RSC plays important roles in maintaining NDRs near promoter regions (Krietenstein et al, 2016;Kubik et al, 2015;Kubik et al, 2017;Musladin et al, 2014), and has been implicated in promoting transcription (Carey et al, 2006;Ganguli et al, 2014;Ocampo et al, 2019;Rawal et al, 2018;Spain et al, 2014;Spain and Govind, 2011). The ability to slide nucleosome to widen NDRs appears to enhance PIC assembly and the TSS selection (Klein-Brill et al, 2019;Kubik et al, 2019).…”

“…To understand the extent to which these defects are caused by changes in chromatin structure, we analyzed MNase-seq data from WT and rsc8 cells (Ocampo et al, 2019). Nucleosomes in both Pol II-B10 and RSC Pro-targets genes moved towards the NDRs in rsc8 cells, leading to shallower NDRs ( Figure 4F).…”