Project description:Human BET family members are promising targets in the therapy of cancer and immunoinflammatory diseases, but their mechanism of action and functional redundancies are poorly understood. Yeast BET factors Bdf1/2 were previously proposed to act as anchors for coactivator TFIID. We investigated their genome wide roles in transcription and found that, while they cooperate with TFIID at many genes, their contributions to transcription are often significantly different. Bdf1/2 co-occupy the majority of yeast promoters and affect preinitiation complex formation by participating in recruitment of TFIID, Mediator and basal factors to chromatin. Surprisingly, we discovered that hypersensitivity of genes to Bdf1/2 depletion results from combined defects in initiation of transcription and early elongation. Bdf1/2 are critical components of yeast transcriptional machinery with many functional similarities to human BET proteins, most notably Brd4.
Project description:Human BET family members are promising targets in the therapy of cancer and immunoinflammatory diseases, but their mechanism of action and functional redundancies are poorly understood. Yeast BET factors Bdf1/2 were previously proposed to act as anchors for coactivator TFIID. We investigated their genome wide roles in transcription and found that, while they cooperate with TFIID at many genes, their contributions to transcription are often significantly different. Bdf1/2 co-occupy the majority of yeast promoters and affect preinitiation complex formation by participating in recruitment of TFIID, Mediator and basal factors to chromatin. Surprisingly, we discovered that hypersensitivity of genes to Bdf1/2 depletion results from combined defects in initiation of transcription and early elongation. Bdf1/2 are critical components of yeast transcriptional machinery with many functional similarities to human BET proteins, most notably Brd4.
Project description:Human BET family members are promising targets in the therapy of cancer and immunoinflammatory diseases, but their mechanism of action and functional redundancies are poorly understood. Yeast BET factors Bdf1/2 were previously proposed to act as anchors for coactivator TFIID. We investigated their genome wide roles in transcription and found that, while they cooperate with TFIID at many genes, their contributions to transcription are often significantly different. Bdf1/2 co-occupy the majority of yeast promoters and affect preinitiation complex formation by participating in recruitment of TFIID, Mediator and basal factors to chromatin. Surprisingly, we discovered that hypersensitivity of genes to Bdf1/2 depletion results from combined defects in initiation of transcription and early elongation. Bdf1/2 are critical components of yeast transcriptional machinery with many functional similarities to human BET proteins, most notably Brd4.
Project description:We report the application of single-molecule-based sequencing technology for high-throughput profiling of histone modifications in yeast. By obtaining bases of sequence from chromatin immunoprecipitated DNA, we generated genome-wide chromatin-state maps of saccharomuces cerevisiae.We find that H3T11 phosphorylationlysine is widely distributed in gene promoter region and chromosome telomere region
Project description:The structural complexity of nucleosomes underlies their functional versatility. Here we report a new type of complexity – nucleosome fragility, manifested as high sensitivity to micrococcal nuclease, in contrast to the common presumption that nucleosomes are similar in resistance to MNase digestion. Using differential MNase digestion of chromatin and high-throughput sequencing, we have identified a special group of nucleosomes termed fragile nucleosomes throughout the yeast genome, nearly one thousand of which are at previously determined “nucleosome free” loci. Nucleosome fragility is broadly implicated in multiple chromatin processes, including transcription, translocation and replication, in correspondence to specific physiological states of cells. In the environmental-stress-response genes, the presence of fragile nucleosomes prior to the occurrence of environmental changes suggests that nucleosome fragility poises genes for swift up-regulation in response to the environmental changes. We propose that nucleosome fragility underscores distinct functional statuses of the chromatin and provides a new dimension for portraying the landscape of genome organization.
Project description:Abf1 and Reb1, two general regulatory factors playing roles at promoters and other genome functional sites in budding yeast, were mapped genome-wide by ChIP-sequencing using strains expressing TAP-tagged versions of the proteins. As expected on the basis of previous in silico analysis of promoter regions, we found that these factors are enriched at the promoters of ribosome biogenesis (Ribi) genes, a large regulon of more than 200 genes required for ribosome biosynthesis and assembly, and known to be coordinately regulated in response to nutrient availability and cellular growth rate.
Project description:The vast landscape of RNA-protein interactions at the heart of post-transcriptional regulation remains largely unexplored. Indeed it is likely that, even in yeast, a substantial fraction of the regulatory RNA-binding proteins (RBPs) remain to be discovered. Systematic experimental methods can play a key role in discovering these RBPs - most of the known yeast RBPs lack RNA-binding domains that might enable this activity to be predicted. We describe here a new proteome-wide approach to identify RNA-protein interactions based on in vitro binding of RNA samples to yeast protein microarrays that represent over 80% of the yeast proteome. We used this procedure to screen for novel RBPs and RNA-protein interactions. A complementary mass spectrometry technique also identified proteins that associate with yeast mRNAs. Both the protein microarray and mass spectrometry methods successfully identify previously annotated RBPs, suggesting that other proteins identified in these assays might be novel RBPs. Of 35 putative novel RBPs identified by either or both of these methods, 12, including 75% of the eight most highly-ranked candidates, reproducibly associated with specific cellular RNAs. Surprisingly, most of the 12 newly discovered RBPs were enzymes. Functional characteristics of the RNA targets of some of the novel RBPs suggest coordinated post-transcriptional regulation of subunits of protein complexes and a possible link between mRNA trafficking and vesicle transport. Our results suggest that many more RBPs still remain to be identified and provide a set of candidates for further investigation.