Virtually all genes in higher organisms are interrupted by regions of noncoding DNA called introns. These introns must be removed from messenger RNA precursors to allow proper gene expression. The process of pre-mRNA splicing is responsible for accurately identifying introns and catalyzing their excision. A fundamental challenge in modern molecular biology is to understand how the splicing apparatus specifically recognizes and removes introns. Because the splicing machinery is highly conserved throughout evolution, my laboratory approaches this question using the genetic and biochemical tools available uniquely in the yeast Saccharomyces cerevisiae.
The RNA-protein particle called the U1 snRNP is the first splicing factor to contact the pre-mRNA, and therefore is a key determinant of splicing specificity. After binding the pre-mRNA, the U1 snRNP proteins interact with other RNAs and proteins to form a bridge that is thought to bring the ends of the intron together for splicing. Therefore, we are very interested in defining the RNAs and proteins that interact with the U1 snRNP.
Currently, we are focusing on the U1 snRNP protein called Prp40. This splicing factor is essential for the splicing reaction and acts at an early step in the splicing pathway. Intriguingly, Prp40 contains conserved amino acid domains of the WWP and dsRNA motifs. Found in a family of proteins including dystrophin, the WWP motif is thought to be a protein-protein interaction domain. In contrast, the dsRNA motif binds double stranded RNA. Thus, Prp40 appears to play a central role in early splicing events. Indeed, we have recently shown that Prp40 interacts directly with other splicing factors early during the splicing pathway. We are now using genetic and biochemical techniques to define these interactions, as this information will shed important new light on how the splicing reaction achieves its high specificity.
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