Our long range goal is to contribute to an understanding of the biochemical mechanisms underlying the regulation of eukaryotic messenger RNA synthesis. The major goal of the research described in this proposal is to determine, at the molecular level, how promoter-specific transcription by RNA polymerase II (Pol II) is controlled by the action of a set of transcription factors that direct accurate initiation and efficient elongation of transcripts in vitro. Our overall approach is essentially the same as that described in the previous proposals: (i) to reconstitute in vitro, with purified proteins, faithful and efficient transcription of eukaryotic protein-coding genes and (ii) to use this reconstituted system to elucidate the biochemical mechanisms governing transcription initiation and elongation by mammalian Pol II. This research will be organized along the following lines: (1) Further analysis of the mechanism of action of Pol II elongation factor Elongin A and of its newly described role in UV-induced ubiquitination of Pol II. (2) Further investigation of the mechanism of action of Pol II elongation factor ELL. (3) Detailed analysis of the mechanisms of action of two novel human ATP-dependent chromatin remodelers referred to as the human INO80 complex and the SNF2 family member and oncogene ALC1 (Activated in Liver Cancer 1). These studies should provide information on the basic mechanisms governing eukaryotic messenger RNA synthesis, and they also have the potential to yield new insights into the molecular bases of cancers resulting from abherrant expression of ELL and its associated proteins and of ALC1.
It is well established that eukaryotic messenger RNA synthesis is a major site for the regulation of gene expression and, further, that its misregulation often results in human diseases, including many forms of cancer. Eukaryotic messenger RNA synthesis is a complex biochemical process catalyzed by the enzyme RNA polymerase II with the assistance of a diverse collection of transcription factors. Despite considerable progress over the past 20 years, critical aspects of the mechanisms underlying misregulation of eukaryotic messenger RNA synthesis in human disease are not completely understood. As a consequence, the goal of the research described in this proposal is to contribute to an understanding of the biochemical mechanisms underlying the regulation of messenger RNA synthesis in mammalian cells. In particular, we plan to pursue investigations of the mechanisms of action of several transcription factors discovered in our lab. In addition to shedding light on the basic mechanisms governing mammalian messenger RNA synthesis, these studies have the potential to shed light on aspects of cancer, since two transcription factors, ELL and ALC1, which we plan to investigate have been proposed to participate in the etiology of acute myeloid leukemia and hepatocellular carcinoma, respectively.
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