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

ABSTRACT: RSC (Remodels the Structure of Chromatin) is a conserved ATP-dependent chromatin remodeling complex that regulates many biological processes, including transcription by RNA polymerase II (Pol II). We report that RSC contributes in generating accessible nucleosomes in transcribed coding sequences (CDSs). RSC MNase ChIP-seq data revealed that RSC-bound nucleosome fragments were very heterogenous (∼80 bp to 180 bp) compared to a sharper profile displayed by the MNase inputs (140 bp to 160 bp), supporting the idea that RSC promotes accessibility of nucleosomal DNA. Notably, RSC binding to +1 nucleosomes and CDSs, but not with -1 nucleosomes, strongly correlated with Pol II occupancies, suggesting that RSC enrichment in CDSs is linked to transcription. We also observed that Pol II associates with nucleosomes throughout transcribed CDSs, and similar to RSC, Pol II-protected fragments were highly heterogenous, consistent with the idea that Pol II interacts with remodeled nucleosomes in CDSs. This idea is supported by the observation that the genes harboring high-levels of Pol II in their CDSs were the most strongly affected by ablating RSC function. Additionally, rapid nuclear depletion of Sth1 decreases nucleosome accessibility and results in accumulation of Pol II in highly transcribed CDSs. This is consistent with a slower clearance of elongating Pol II in cells with reduced RSC function, and is distinct from the effect of RSC depletion on PIC assembly. Altogether, our data provide evidence in support of the role of RSC in promoting Pol II elongation, in addition to its role in regulating transcription initiation.

SUBMITTER: Biernat E

PROVIDER: S-EPMC8049546 | biostudies-literature |

REPOSITORIES: biostudies-literature

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

Project description:RSC (Remodels the Structure of Chromatin) is a conserved ATP-dependent chromatin remodeling complex that regulates many biological processes, including transcription by RNA polymerase II (Pol II). We report that not only RSC binds to nucleosomes in coding sequences (CDSs) but also remodels them to promote transcription. RSC MNase ChIP-seq data revealed that RSC-protected fragments were very heterogenous (~80 bp to 180 bp) compared to the sharper profile displayed by the MNase inputs (140 bp to 160 bp), supporting the idea that RSC activity promotes accessibility of nucleosomal DNA. Importantly, RSC binding to +1 nucleosomes and CDSs, but not with -1 nucleosomes, strongly correlated with Pol II occupancies suggesting that the RSC enrichment in CDSs is important for efficient transcription. This is further supported by a similar heterogenous distribution of Pol II-protected fragments. As such, the genes harboring high-levels of RSC in their CDSs were the most strongly affected by ablating RSC function. Altogether, this study provides a mechanism by which RSC-mediated remodeling aids in RNA Pol II traversal though coding sequence nucleosomes in vivo.

2021-04-21 | GSE147065 | GEO

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

Project description:The RSC complex remodels nucleosomes in transcribed coding sequences and promotes transcription in Saccharomyces cerevisiae

| PRJNA612872 | ENA

Recruitment of Saccharomyces cerevisiae Cmr1/Ydl156w to Coding Regions Promotes Transcription Genome Wide.

Project description:Cmr1 (changed mutation rate 1) is a largely uncharacterized nuclear protein that has recently emerged in several global genetic interaction and protein localization studies. It clusters with proteins involved in DNA damage and replication stress response, suggesting a role in maintaining genome integrity. Under conditions of proteasome inhibition or replication stress, this protein localizes to distinct sub-nuclear foci termed as intranuclear quality control (INQ) compartments, which sequester proteins for their subsequent degradation. Interestingly, it also interacts with histones, chromatin remodelers and modifiers, as well as with proteins involved in transcription including subunits of RNA Pol I and Pol III, but not with those of Pol II. It is not known whether Cmr1 plays a role in regulating transcription of Pol II target genes. Here, we show that Cmr1 is recruited to the coding regions of transcribed genes of S. cerevisiae. Cmr1 occupancy correlates with the Pol II occupancy genome-wide, indicating that it is recruited to coding sequences in a transcription-dependent manner. Cmr1-enriched genes include Gcn4 targets and ribosomal protein genes. Furthermore, our results show that Cmr1 recruitment to coding sequences is stimulated by Pol II CTD kinase, Kin28, and the histone deacetylases, Rpd3 and Hos2. Finally, our genome-wide analyses implicate Cmr1 in regulating Pol II occupancy at transcribed coding sequences. However, it is dispensable for maintaining co-transcriptional histone occupancy and histone modification (acetylation and methylation). Collectively, our results show that Cmr1 facilitates transcription by directly engaging with transcribed coding regions.

| S-EPMC4744024 | biostudies-literature

Chromatin remodelling complex RSC promotes base excision repair in chromatin of Saccharomyces cerevisiae.

Project description:The base excision repair (BER) pathway is a conserved DNA repair system required to maintain genomic integrity and prevent mutagenesis in all eukaryotic cells. Nevertheless, how BER operates in vivo (i.e. in the context of chromatin) is poorly understood. We have investigated the role of an essential ATP-dependent chromatin remodelling (ACR) complex RSC (Remodels the Structure of Chromatin) in BER of intact yeast cells. We show that depletion of STH1, the ATPase subunit of RSC, causes enhanced sensitivity to the DNA alkylating agent methyl methanesulfonate (MMS) and results in a substantial inhibition of BER, at the GAL1 locus and in the genome overall. Consistent with this observation, the DNA in chromatin is less accessible to micrococcal nuclease digestion in the absence of RSC. Quantitative PCR results indicate that repair deficiency in STH1 depleted cells is not due to changes in the expression of BER genes. Collectively, our data indicates the RSC complex promotes efficient BER in chromatin. These results provide, for the first time, a link between ATP-dependent chromatin remodelling and BER in living cells.

| S-EPMC4026264 | biostudies-literature

Transcription Promotes the Interaction of the FAcilitates Chromatin Transactions (FACT) Complex with Nucleosomes in <i>Saccharomyces cerevisiae</i>.

Project description:The FACT (FAcilitates Chromatin Transactions) complex is a conserved complex that maintains chromatin structure on transcriptionally active genes. Consistent with this, FACT is enriched on highly expressed genes, but how it is targeted to these regions is unknown. In vitro, FACT binds destabilized nucleosomes, supporting the hypothesis that FACT is targeted to transcribed chromatin through recognition of RNA polymerase (RNAP)-disrupted nucleosomes. In this study, we used high-resolution analysis of FACT occupancy in Saccharomyces cerevisiae to test this hypothesis. We demonstrate that FACT interacts with nucleosomes in vivo and that its interaction with chromatin is dependent on transcription by any of the three RNAPs. Deep sequencing of micrococcal nuclease-resistant fragments shows that FACT-bound nucleosomes exhibit differing nuclease sensitivity compared to bulk chromatin, consistent with a modified nucleosome structure being the preferred ligand for this complex. Interestingly, a subset of FACT-bound nucleosomes may be "overlapping dinucleosomes," in which one histone octamer invades the ∼147-bp territory normally occupied by the adjacent nucleosome. While the differing nuclease sensitivity of FACT-bound nucleosomes could also be explained by the demonstrated ability of FACT to alter nucleosome structure, transcription inhibition restores nuclease resistance, suggesting that it is not due to FACT interaction alone. Collectively, these results are consistent with a model in which FACT is targeted to transcribed genes through preferential interaction with RNAP-disrupted nucleosomes.

| S-EPMC6218215 | biostudies-literature

RSC Chromatin-Remodeling Complex Is Important for Mitochondrial Function in Saccharomyces cerevisiae.

Project description:RSC (Remodel the Structure of Chromatin) is an ATP-dependent chromatin remodeling complex essential for the growth of Saccharomyces cerevisiae. RSC exists as two distinct isoforms that share core subunits including the ATPase subunit Nps1/Sth1 but contain either Rsc1or Rsc2. Using the synthetic genetic array (SGA) of the non-essential null mutation method, we screened for mutations exhibiting synthetic growth defects in combination with the temperature-sensitive mutant, nps1-105, and found connections between mitochondrial function and RSC. rsc mutants, including rsc1Δ, rsc2Δ, and nps1-13, another temperature-sensitive nps1 mutant, exhibited defective respiratory growth; in addition, rsc2Δ and nps1-13 contained aggregated mitochondria. The rsc2Δ phenotypes were relieved by RSC1 overexpression, indicating that the isoforms play a redundant role in respiratory growth. Genome-wide expression analysis in nps1-13 under respiratory conditions suggested that RSC regulates the transcription of some target genes of the HAP complex, a transcriptional activator of respiratory gene expression. Nps1 physically interacted with Hap4, the transcriptional activator moiety of the HAP complex, and overexpression of HAP4 alleviated respiratory defects in nps1-13, suggesting that RSC plays pivotal roles in mitochondrial gene expression and shares a set of target genes with the HAP complex.

| S-EPMC4472808 | biostudies-literature

Protein kinase CK2 holoenzyme promotes start-specific transcription in Saccharomyces cerevisiae.

Project description:In Saccharomyces cerevisiae, the entrance into S phase requires the activation of a specific burst of transcription, which depends on SBF (SCB binding factor, Swi4/Swi6) and MBF (MCB binding factor, Mbp1/Swi6) complexes. CK2 is a pleiotropic kinase involved in several cellular processes, including the regulation of the cell cycle. CK2 is composed of two catalytic subunits (? and ?') and two regulatory subunits (? and ?'), both of which are required to form the active holoenzyme. Here we investigate the function of the CK2 holoenzyme in Start-specific transcription. The ckb1? ckb2? mutant strain, bearing deletions of both genes encoding CK2 regulatory subunits, shows a delay of S-phase entrance due to a severe reduction of the expression of SBF- and MBF-dependent genes. This transcriptional defect is caused by an impaired recruitment of Swi6 and Swi4 to G1 gene promoters. Moreover, CK2 ? and ?' subunits interact with RNA polymerase II, whose binding to G1 promoters is positively regulated by the CK2 holoenzyme. Collectively, these findings suggest a novel role for the CK2 holoenzyme in the activation of G1 transcription.

| S-EPMC3811569 | biostudies-literature

Evidence for abundant transcription of non-coding regions in the Saccharomyces cerevisiae genome.

Project description:BackgroundRecent studies in a growing number of organisms have yielded accumulating evidence that a significant portion of the non-coding region in the genome is transcribed. We address this issue in the yeast Saccharomyces cerevisiae.ResultsTaking into account the absence of a significantly large yeast EST database, we use microarray expression data collected for genomic regions erroneously believed to be coding to study the expression pattern of non-coding regions in the Saccharomyces cerevisiae genome. We find that at least 164 out of 589 (28%) such regions are expressed under specific biological conditions. In particular, looking at the probes that are located opposing other known genes at the same genomic locus, we find that 88 out of 341 (26%) of these genes support antisense transcription. The expression patterns of these antisense genes are positively correlated. We validate these results using RT-PCR on a sample of 6 non-coding transcripts.Conclusion1. The yeast genome is transcribed on a scale larger than previously assumed. 2. Correlated transcription of antisense genes is abundant in the yeast genome. 3. Antisense genes in yeast are non-coding.

| S-EPMC1181813 | biostudies-literature

Chromatin- and transcription-related factors repress transcription from within coding regions throughout the Saccharomyces cerevisiae genome.

Project description:Previous studies in Saccharomyces cerevisiae have demonstrated that cryptic promoters within coding regions activate transcription in particular mutants. We have performed a comprehensive analysis of cryptic transcription in order to identify factors that normally repress cryptic promoters, to determine the amount of cryptic transcription genome-wide, and to study the potential for expression of genetic information by cryptic transcription. Our results show that a large number of factors that control chromatin structure and transcription are required to repress cryptic transcription from at least 1,000 locations across the S. cerevisiae genome. Two results suggest that some cryptic transcripts are translated. First, as expected, many cryptic transcripts contain an ATG and an open reading frame of at least 100 codons. Second, several cryptic transcripts are translated into proteins. Furthermore, a subset of cryptic transcripts tested is transiently induced in wild-type cells following a nutritional shift, suggesting a possible physiological role in response to a change in growth conditions. Taken together, our results demonstrate that, during normal growth, the global integrity of gene expression is maintained by a wide range of factors and suggest that, under altered genetic or physiological conditions, the expression of alternative genetic information may occur.

| S-EPMC2581627 | biostudies-literature

Transcribed germline-limited coding sequences in Oxytricha trifallax.

Project description:The germline-soma divide is a fundamental distinction in developmental biology, and different genes are expressed in germline and somatic cells throughout metazoan life cycles. Ciliates, a group of microbial eukaryotes, exhibit germline-somatic nuclear dimorphism within a single cell with two different genomes. The ciliate Oxytricha trifallax undergoes massive RNA-guided DNA elimination and genome rearrangement to produce a new somatic macronucleus (MAC) from a copy of the germline micronucleus (MIC). This process eliminates noncoding DNA sequences that interrupt genes and also deletes hundreds of germline-limited open reading frames (ORFs) that are transcribed during genome rearrangement. Here, we update the set of transcribed germline-limited ORFs (TGLOs) in O. trifallax. We show that TGLOs tend to be expressed during nuclear development and then are absent from the somatic MAC. We also demonstrate that exposure to synthetic RNA can reprogram TGLO retention in the somatic MAC and that TGLO retention leads to transcription outside the normal developmental program. These data suggest that TGLOs represent a group of developmentally regulated protein-coding sequences whose gene expression is terminated by DNA elimination.

| S-EPMC8495736 | biostudies-literature

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