Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

H2A.Z (pht1) ChIP-seq data in 968, Y0036, and swc5-del strains

ABSTRACT: After finding a QTL hotspot in a fission yeast cross of strains 968xY0036 that was apparently driven by frame shift mutation in swc5 (swc5-fs), we have determined the genome-wide occupancy of the histone variant H2AZ (PhT1). Indeed swc5 has been shown to affect H2AZ occupancy. To test the impact of swc5-fs on H2A.Z deposition, we performed genome-wide chromatin immuno-precipitation of H2A.Z coupled with deep sequencing (ChIP-seq) in the two parental strains (968, Y0036) and in a swc5 deletion strain. We also assed the histone H3 occupancy as a positive control. We generated pht1-myc tagged strains to precipitate H2AZ with Myc antibody. Several negative controls have been performed, in particular pulldowns with the non tagged strains. Results show a reduced H2A.Z occupancy at the +1 histone in both Y0036 and swc5-deletion strains.

INSTRUMENT(S): Illumina MiSeq

ORGANISM(S): Schizosaccharomyces pombe

SUBMITTER: Mathieu Clement-Ziza

PROVIDER: E-MTAB-2650 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Natural genetic variation impacts expression levels of coding, non-coding, and antisense transcripts in fission yeast.

Clément-Ziza Mathieu M Marsellach Francesc X FX Codlin Sandra S Papadakis Manos A MA Reinhardt Susanne S Rodríguez-López María M Martin Stuart S Marguerat Samuel S Schmidt Alexander A Lee Eunhye E Workman Christopher T CT Bähler Jürg J Beyer Andreas A

Molecular systems biology 20141128

Our current understanding of how natural genetic variation affects gene expression beyond well-annotated coding genes is still limited. The use of deep sequencing technologies for the study of expression quantitative trait loci (eQTLs) has the potential to close this gap. Here, we generated the first recombinant strain library for fission yeast and conducted an RNA-seq-based QTL study of the coding, non-coding, and antisense transcriptomes. We show that the frequency of distal effects (trans-eQT ...[more]

PMID: 25432776

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Project description:H2A.Z is a highly conserved histone variant involved in several key nuclear processes. It is incorporated into promoters by SWR-C-related chromatin remodeling complexes, but whether it is also actively excluded from non-promoter regions is not clear. Here, we provide genomic and biochemical evidence that RNA polymerase II (RNAPII) elongation-associated histone chaperones FACT and Spt6 both contribute to restricting H2A.Z from intragenic regions. In the absence of FACT or Spt6, the lack of H2A.Z eviction, coupled to its pervasive incorporation by mislocalized SWR-C, alters chromatin composition and contributes to cryptic initiation. Thus, chaperone-mediated H2A.Z removal is crucial for restricting the chromatin signature of gene promoters, which otherwise may license or promote cryptic transcription. We profiled H2A.Z occupancy in S. cerevisiae by ChIP-chip on tiling arrays in different mutants for chromatin remodelers and histone chaperones. In most experiments, H2A.Z ChIP samples (Cy5-labeled) were performed using a polyclonal antibody against H2A.Z and hybridized against H2B ChIP samples (Cy3-labeled), also performed using a polyclonal antibody. Temperature-sensitive mutants for the histone chaperones Spt16 and Spt6 (spt16-197 and spt6-1004 respectively) showed strong H2A.Z localization defects so the role of these factors in H2A.Z localization was further analyzed. H2A.Z ChIP-chip experiments in spt16-197 and spt6-1004 were repeated using different experimental designs [normalized against Input DNA or Mock (IgG) ChIP samples]. The contribution of Spt16 and Spt6 on H2A.Z localization was also confirmed by nuclear depletion of Spt16 and Spt6 using the anchor-away system. H2A.Z ChIP-chip experiments were also performed in sic1Δ and spt16-197/sic1Δ cells in order to rule out any G1-arrest artifact. H2A.Z ChIP-chip experiments were repeated using spike-in controls for normalization, revealing widespread H2A.Z occupancy in Spt16 and Spt6 mutants. In addition, the localization of the chromatin remodeler SWR-C was determined in wild type cells as well as in spt16-197 and spt6-1004 cells. Finally, we also profiled histone H4 occupancy by ChIP-chip in wild type, spt16-197, spt6-1004, spt16-197/htz1Δ and spt6-1004/htz1Δ cells. All ChIP-chip experiments were done in duplicates. Each microarray was normalized using the Lima Loess and replicates were combined using a weighted average method as previously described (Pokholok et al., 2005).

Dataset Information

H2A.Z (pht1) ChIP-seq data in 968, Y0036, and swc5-del strains

Publications

Natural genetic variation impacts expression levels of coding, non-coding, and antisense transcripts in fission yeast.

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