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A Role for FACT in RNA Polymerase II Promoter-Proximal Pausing

ABSTRACT: This SuperSeries is composed of the SubSeries listed below. FACT (Facilitates Chromatin Transcription) is an evolutionarily conserved histone chaperone that was initially identified as an activity capable of promoting RNA polymerase II transcription through nucleosomes in vitro. In this report, we describe a global analysis of FACT function in Pol II transcription in Drosophila. We present evidence that loss of FACT has a dramatic impact on Pol II elongation-coupled processes including H3K4 and H3K36 methylation, consistent with a role for FACT in coordinating histone modification and chromatin architecture during Pol II transcription. Importantly, we identify a role for FACT in the maintenance of promoter proximal Pol II pausing, a key step in transcription activation in higher eukaryotes. These findings bring to light a broader role for FACT in the regulation of Pol II transcription.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE129236 | GEO | 2019/06/14

REPOSITORIES: GEO

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Project description:RSC (remodels the structure of chromatin) is an essential ATP-dependent chromatin remodeling complex in Saccharomyces cerevisiae. The catalytic subunit of RSC, Sth1 uses its ATPase activity to slide or remove nucleosomes. RSC has been shown to regulate the width of the nucleosome-depleted regions (NDRs) by sliding the flanking nucleosomes away from NDRs. As such the nucleosomes encroach NDRs when RSC is depleted and leads to transcription initiation defects. In this study, we examined the effects of the catalytic-dead Sth1 on transcription and compared them to the effects observed during acute and rapid Sth1 depletion by auxin-induced degron strategy. We found that rapid depletion of Sth1 reduces recruitment of TBP and Pol II in highly transcribed genes, as would be expected considering its role in regulating chromatin structure at promoters. In contrast, cells harboring the catalytic-dead Sth1 exhibited a severe reduction in TBP binding, but surprisingly, also displayed a substantial accumulation in Pol II occupancies within coding regions. After depleting endogenous Sth1 in the catalytic dead mutant, we observed a further increase in Pol II occupancies, suggesting that the inactive Sth1 contributed to the observed accumulation of Pol II in coding regions. Notwithstanding the Pol II increase, the ORF occupancies of histone chaperones FACT and Spt6 were significantly reduced in the mutant. These results suggest a potential role for RSC in recruiting/retaining these chaperones in coding regions. Pol II accumulation despite substantial reductions in TBP, FACT, and Spt6 occupancies in the catalytic-dead mutant could be indicative of severe transcription elongation and termination defects. Such defects would be consistent with studies showing that RSC is recruited to coding regions in a transcription-dependent manner. Thus, these findings imply a role for RSC in transcription elongation and termination processes, in addition to its established role in transcription initiation.

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A Role for FACT in RNA Polymerase II Promoter-Proximal Pausing

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