Project description:ChIP-chip assays to determine the localisation of 3xFLAG tagged Isw1 in wild-type yeast. Two color experiment. ChIP/Input. Biological replicates=3. Other data used in this study are provided in GEO Series GSE31015 and GSE31833.
Project description:Aberrant forms of the SWI/SNF chromatin remodeling complex are associated with human disease. Loss of the Snf5 subunit of SWI/SNF is a driver mutation in pediatric rhabdoid cancers and forms aberrant sub-complexes that are not well characterized. We determined the effects of loss of Snf5 on the composition, nucleosome binding, recruitment and remodeling activities of yeast SWI/SNF. The Snf5 subunit interacts with the ATPase domain of Snf2 and forms a submodule consisting of Snf5, Swp82 and Taf14 as shown by mapping SWI/SNF subunit interactions by crosslinking-mass spectrometry and subunit deletion followed by immunoaffinity chromatography. Snf5 promoted binding of the Snf2 ATPase domain to nucleosomal DNA, enhanced its catalytic activity and facilitated nucleosome remodeling. Snf5 was required for acidic transcription factors to recruit SWI/SNF to chromatin. RNA-seq analysis suggested that both the recruitment and catalytic functions mediated by Snf5 are required for SWI/SNF regulation of gene expression.
Project description:We report the genomic localization of cohesin oligomers in nocodazole arrested yeast cells. Two alleles of SMC3 were expressed in yeast cells, one fused to BirA enzyme and the other tagged with AviTag. Cohesin oligomers were biotinylated and ChIP with streptavidin beads. As control experiments, cohesin localization on chromosome was determined in strains expresses freely diffusable BirA enzyme, where all Smc3 proteins were biotinylated; non-specific ChIP were determined in strains with no BirA.
Project description:We report the application of single-molecule-based sequencing technology for high-throughput profiling of SNF5 binding in human pluripotent embryonic carcinoma NCCIT and SNF5 overexpressed NCCIT cells. We generated genome-wide cSNF5 maps of NCCIT and SNF5 overexpressed NCCIT cells from chromatin immunoprecipitated DNA. SNF5 and OCT4 seem not to share their binding in OCT4 centered binding plot in control, while SNF5 overexpression directs SNF5 to OCT4 target genes.
Project description:Epigenetic alterations have been increasingly implicated in oncogenesis. Analysis of Drosophila mutants suggests that Polycomb and SWI/SNF complexes can serve antagonistic developmental roles. However, the relevance of this relationship to human disease is unclear. Here we have investigated functional relationships between these epigenetic regulators in oncogenic transformation. Mechanistically, we show that loss of the SNF5 tumor suppressor leads to elevated expression of the Polycomb gene EZH2 and that Polycomb targets are broadly H3K27-trimethylated and repressed in SNF5-deficient fibroblasts and cancers. Further, we show antagonism between SNF5 and EZH2 in the regulation of stem cell-associated programs and that Snf5 loss activates those programs. Finally, using conditional mouse models, we show that inactivation of Ezh2 blocks tumor formation driven by Snf5 loss.
Project description:We report the application of single-molecule-based sequencing technology for high-throughput profiling of SNF5 binding in human pluripotent embryonic carcinoma NCCIT and SNF5 overexpressed NCCIT cells. We generated genome-wide cSNF5 maps of NCCIT and SNF5 overexpressed NCCIT cells from chromatin immunoprecipitated DNA. SNF5 and OCT4 seem not to share their binding in OCT4 centered binding plot in control, while SNF5 overexpression directs SNF5 to OCT4 target genes. Examination of the relationship between SNF5 and OCT4 binding in control and SNF5 overexpressed cells.