Mechanisms that Couple Chromatin Modifications to Transcription

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Project Summary The goal of this research program is to elucidate the proteins and mechanisms that regulate transcription within the context of chromatin, a process critical to every eukaryotic cell. Nucleosomes pose barriers to RNA polymerase II (Pol II) that must be overcome for accurate and efficient gene expression, and whose properties and interactions are modulated by post-translational modification. During transcription elongation, a conserved set of factors assembles with Pol II and facilitates its transit by altering the stability, positioning, and post- translational modification states of nucleosomes. The proposal addresses three major challenges related to these functions of eukaryotic transcription elongation factors. (1) What are the mechanisms by which transcription elongation factors couple chromatin changes, including histone modifications, to RNA synthesis? (2) How are the patterns of these epigenetic modifications determined? (3) What are the primary versus indirect functions of core components of the Pol II elongation machinery? These questions will be approached through a comprehensive analysis of the Paf1 complex (Paf1C) and proteins with which it interacts. Paf1C is a highly conserved transcription elongation factor that globally associates with Pol II on the bodies of active genes. The multifunctional nature of Paf1C affords a unique opportunity to reveal how transcription is coupled to co-transcriptional events. To this end, a multifaceted approach comprising innovative genetic and proteomic screens, mechanistic biochemistry, and genomics will be deployed. This project will determine how Paf1C stimulates critical histone modifications and interfaces with a chromatin remodeling factor with genetic links to prostate cancer. In-depth studies of the interactions between Paf1C and the Pol II elongation complex will uncover the molecular mechanisms that spatially constrain Paf1C-dependent chromatin changes to active genes. Finally, the primary and subunit-specific functions of Paf1C, as well as the cellular pathways that compensate for its absence, will be determined. The studies will be performed in Saccharomyces cerevisiae to capitalize on the sophisticated tools developed for that system. Given the strong conservation of all proteins and histone modifications studied, the conceptual advances that arise from this work will have direct implications for the understanding of gene regulation in humans, where defects in this process cause a wide range of cancers and other diseases.