| The highly conserved Cap Binding Complex (CBC) is comprised of two subunits, Cbp20 and Cbp80, and binds to the 5' end of pre-mRNA as it emerges from the transcribing RNA polymerase (PolII). Evidence suggesting a role for the CBC in multiple reactions during gene expression, including transcription, splicing, 3' end processing, and RNA export, prompted us to hypothesize that the CBC is involved in coupling these reactions, and that this coordinated regulation facilitates the ability of a cell to respond to stress. The research presented in this dissertation specifically dissects the roles of the CBC in splicing and in transcription and provides insight into a mechanism by which the CBC might contribute to the coupling of these two reactions in Saccharomyces cerevisiae. We have demonstrated that the CBC is specifically required for the splicing of two intron-containing genes with a particular non-consensus 5' splice site, SUS1 and GCR1. Through its effects on splicing, we found that the CBC contributes to the regulation of a histone modification and the acitivity of a key glycolytic transcription factor, resulting in genome-wide changes to gene expression when the CBC is deleted. We discovered that the CBC is down-regulated at multiple levels in response to nutrient stress, suggesting that the coordinated activities of the CBC are particularly important under these conditions. One of these levels of regulation appears to be proteolysis by the proteasome. Furthermore, a role for the CBC in repressing expression of Rpn4, a critical regulator of proteasome levels, suggests that the CBC is involved in a feedback loop in which its targeting to the proteasome is determined by its ability to influence gene expression. Directed mutagenesis of evolutionarily-conserved domains of Cbp80 allowed us to uncouple the functions of the CBC in gene expression revealing a role for the CBC in transcription that is independent of its ability to act in splicing. In sum, the findings presented here demystify the role of the CBC in gene expression by enhancing our understanding of the CBC's role in splicing, demonstrating novel functions for the CBC in transcription, identifying multiple mechanisms by which the CBC is regulated, and illustrating a requirement for the CBC in the cellular stress response. |
