The RSC chromatin remodeling complex has a crucial role in the complete remodeler set for yeast <i>PHO5</i> promoter opening

Musladin, Sanja; Krietenstein, Nils; Korber, Philipp; Barbarić, Slobodan

doi:10.1093/nar/gkt1395

Cited by 39 publications

(64 citation statements)

References 68 publications

(166 reference statements)

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“…These effects are likely dominant to differences in the physical properties between different histone variants. However, these highly-delayed kinetics – especially at the −1 nucleosome compared to the −2 nucleosome – are slower than those observed when using strains with multiple remodeler deletions (Musladin et al, 2014), which seems inconsistent with incompatibility with chromatin remodelers being the sole explanation. Thus, this argues for additional effects owing to differences in yeast versus human nucleosomes.…”

Section: Discussionmentioning

confidence: 89%

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Resetting the Yeast Epigenome with Human Nucleosomes

Truong

Boeke

2017

Cell

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Summary Humans and yeast are separated by a billion years of evolution, yet their conserved histones retain central roles in gene regulation. Here, we “reset” yeast to use core human nucleosomes in lieu of their own – a rare event taking twenty days – which initially only worked with variant H3.1. The cells adapt by acquiring suppressor mutations in cell-division genes, or by acquiring certain aneuploid states. Converting five histone residues to their yeast counterparts restored robust growth. We reveal that humanized nucleosomes are positioned according to endogenous yeast DNA sequence and chromatin-remodeling network, as judged by a yeast-like nucleosome repeat length. However, human nucleosomes have higher DNA occupancy, globally reduce RNA content, and slow adaptation to new conditions by delaying chromatin remodeling. These humanized yeasts (including H3.3) pose fundamental new questions about how chromatin is linked to many cell processes, and provides a platform to study histone variants via yeast epigenome reprogramming.

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Section: Discussionmentioning

confidence: 89%

“…Importantly, the PHO5 promoter relies on multiple chromatin remodelers (Musladin et al, 2014), and functions independently of replication (Schmid et al, 1992), thus, its remodeling should not be confounded by cell-cycle alterations. As with others, this promoter also features a highly occupied −1 nucleosome in the humanized cells (Figure 6E).…”

Section: Resultsmentioning

confidence: 99%

Resetting the Yeast Epigenome with Human Nucleosomes

Truong

Boeke

2017

Cell

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“…4). Histone acetylation by Gcn5 could stimulate the recruitment or function of a remodeler besides SWI/SNF, as there is evidence that RSC, Isw1, and Chd1 act with SWI/SNF and INO80 in nucleosome remodeling at PHO5 (Musladin et al 2014), and functional overlap among different remodeling complexes occurs in mammalian cells (Morris et al 2014).…”

Section: Wwwgenomeorgmentioning

confidence: 99%

“…Eliminating Snf2, a catalytic subunit of SWI/SNF, reduces nucleosome remodeling at PHO8 (Gregory et al 1999) and impairs remodeling and eviction at PHO5; however, binding of activator Pho4 to the PHO5 UAS is also reduced in snf2Δ cells (Adkins et al 2007;Barbaric et al 2007). Reduced Pho4 binding could also apply to overlapping functions described for SWI/SNF, Ino80, RSC, Isw1, and Chd1 in nucleosome remodeling at PHO5 (Musladin et al 2014). SWI/SNF is needed for efficient nucleosome remodeling (Hirschhorn et al 1992) and eviction at SUC2 (Schwabish and Struhl 2007), GAL1 , HO (Gkikopoulos et al 2009), RNR1 (Sharma et al 2003), and CHA1 ), but whether it functions upstream of or downstream from activator binding is not always clear.…”

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confidence: 99%

Genome-wide cooperation by HAT Gcn5, remodeler SWI/SNF, and chaperone Ydj1 in promoter nucleosome eviction and transcriptional activation

Qiu

Chereji

et al. 2015

Genome Res.

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Chaperones, nucleosome remodeling complexes, and histone acetyltransferases have been implicated in nucleosome disassembly at promoters of particular yeast genes, but whether these cofactors function ubiquitously, as well as the impact of nucleosome eviction on transcription genome-wide, is poorly understood. We used chromatin immunoprecipitation of histone H3 and RNA polymerase II (Pol II) in mutants lacking single or multiple cofactors to address these issues for about 200 genes belonging to the Gcn4 transcriptome, of which about 70 exhibit marked reductions in H3 promoter occupancy on induction by amino acid starvation. Examining four target genes in a panel of mutants indicated that SWI/SNF, Gcn5, the Hsp70 cochaperone Ydj1, and chromatin-associated factor Yta7 are required downstream from Gcn4 binding, whereas Asf1/Rtt109, Nap1, RSC, and H2AZ are dispensable for robust H3 eviction in otherwise wild-type cells. Using ChIP-seq to interrogate all 70 exemplar genes in single, double, and triple mutants implicated Gcn5, Snf2, and Ydj1 in H3 eviction at most, but not all, Gcn4 target promoters, with Gcn5 generally playing the greatest role and Ydj1 the least. Remarkably, these three cofactors cooperate similarly in H3 eviction at virtually all yeast promoters. Defective H3 eviction in cofactor mutants was coupled with reduced Pol II occupancies for the Gcn4 transcriptome and the most highly expressed uninduced genes, but the relative Pol II levels at most genes were unaffected or even elevated. These findings indicate that nucleosome eviction is crucial for robust transcription of highly expressed genes but that other steps in gene activation are more rate-limiting for most other yeast genes.

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“…SWI/SNF could be replaced by RSC (Fig. S6C) (14), whereas addition of the histone chaperones Nap1 and Asf1 was without effect. The promoter specificity of transcription was confirmed by the use of a mutant chromatin template, in which the 24-bp PHO5 core promoter sequence containing the TATA box was replaced by an unrelated sequence (12); virtually no specific transcripts were obtained ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Chromatin potentiates transcription

Nagai

Davis

Mattei

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Chromatin isolated from the chromosomal locus of the PHO5 gene of yeast in a transcriptionally repressed state was transcribed with 12 pure proteins (80 polypeptides): RNA polymerase II, six general transcription factors, TFIIS, the Pho4 gene activator protein, and the SAGA, SWI/SNF, and Mediator complexes. Contrary to expectation, a nucleosome occluding the TATA box and transcription start sites did not impede transcription but rather, enhanced it: the level of chromatin transcription was at least sevenfold greater than that of naked DNA, and chromatin gave patterns of transcription start sites closely similar to those occurring in vivo, whereas naked DNA gave many aberrant transcripts. Both histone acetylation and trimethylation of H3K4 (H3K4me3) were important for chromatin transcription. The nucleosome, long known to serve as a general gene repressor, thus also performs an important positive role in transcription.RNA polymerase II | PHO5 | Saccharomyces cerevisiae A ssembly of purified histones on promoter DNA interferes with the initiation of transcription by RNA polymerase II and general transcription factors (GTFs) in vitro (1) and in vivo (2). Factors that relieve inhibition by histones have been identified by transcribing chromatin reconstituted with purified histones and genetic analysis. These factors include ATP-dependent chromatin remodelers (3, 4), histone-modifying enzymes (5-7), FACT (8), and TFIIS (9). Although informative, these studies are incomplete, because chromatin reconstituted with purified histones differs from chromatin assembled in vivo. Reconstituted chromatin lacks the patterns of histone modification, histone variants, and nonhistone proteins shown to play important roles in transcription in vivo. Nucleosome positioning, also important for transcription in vivo, cannot be accurately reconstituted in vitro. We have, therefore, investigated chromatin assembled in vivo as a template for transcription in vitro. ResultsPHO5 chromatin in the transcriptionally repressed state was excised from yeast chromosomes in circular form by recombination and purified by affinity chromatography as described ( Fig. S1 A and B) (10, 11). PHO5 chromatin isolated in this way was indistinguishable from chromatin at the chromosomal locus on the basis of digestion with specific and nonspecific endonucleases ( Fig. S1 C and D) (10,12). Because the chromatin was derived from a single copy gene, very small quantities were obtained: on the order of 10 fmol/L cell culture. At this low level, transcription cannot be detected directly by radioisotope incorporation or fluorescence, and therefore, we turned to RT-PCR. Two sets of primers were used: one to amplify the region downstream of the transcription start sites (TSSs) and detect all transcripts ("downstream" primer pair) and one to amplify any signal from cryptic transcripts originating upstream and reading through the promoter ("upstream" primer pair) (Fig. 1A and Fig. S2A). Subtraction of the upstream signal from the downstream signal revealed the level of p...

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The RSC chromatin remodeling complex has a crucial role in the complete remodeler set for yeast PHO5 promoter opening

Cited by 39 publications

References 68 publications

Resetting the Yeast Epigenome with Human Nucleosomes

Resetting the Yeast Epigenome with Human Nucleosomes

Genome-wide cooperation by HAT Gcn5, remodeler SWI/SNF, and chaperone Ydj1 in promoter nucleosome eviction and transcriptional activation

Chromatin potentiates transcription

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