The long-term objectives of the proposed project are to elucidate the mechanism of action and in vivo function of MOT1, an essential yeast protein that regulates transcription by dissociating TATA-binding protein (TBP)-DNA complexes in an ATP-dependent manner. MOT1 is a member of a large and growing family of evolutionarily conserved nuclear proteins (the SNF2/SWI2 family) involved in transcription, DNA repair, recombination, and chromosome segregation. Mutation of SNF2/SWI2 family members in humans have been identified as the causes of Cockayne's syndrome, X-linked mental retardation, and alpha-thalassemia. Despite the ubiquitous occurrence of SNF2/SWI2 family members, the molecular mechanisms of action of these proteins are not understood in detail, nor is it understood what roles most of these proteins play in vivo. Biochemical, molecular biological and genetic approaches will be used to define in molecular detail how MOT1 interacts with TBP, and how its ATPase is utilized to drive TBP-DNA complex dissociation. These approaches will also be used to identify other proteins that MOT1 interacts with in vivo and to define the parameters that determine the magnitude of MOT1's effect on the expression of individual genes in vivo. The proposed analysis of MOT1 function will lead to a better understanding of the role of MOT1 as well as a better understanding of the functions of SNF2/SWI2 family members in general. The proposed work will also provide a framework for the analysis of other SNF2/SWI2 family members whose activities are currently poorly understood.
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