Identification of critical residues in spliceosomal protein Dib1 (939.6)

Abstract

Pre‐messenger RNA in eukaryotic cells must have its introns removed in order to be able toproduce functional proteins. This removal of introns is facilitated by a macromolecule known as the spliceosome. The spliceosome is a large complex of five snRNA and ~80 proteins that orients the pre‐mRNA such that two transesterification reactions are catalyzed resulting in the removal of introns from pre‐mRNA resulting in a mature mRNA. The mRNA can then be translated in to a functional protein. Mutations in the spliceosome and the proteins that regulate the spliceosome can have drastic effects and lead to diseases such as Retinitis Pigmentosa. Dib1 is an evolutionarily conserved protein in the spliceosome associated with the U5 snRNP in the spliceosome and is required for cell viability. Although the human homolog, Dim1, has been crystalized its role still remains unknown. In order to identify Dib1’s function, we have created point mutations in the protein and analyzed their effects on cell viability. We have identified a variety of temperature sensitive and lethal dib1 mutants in Saccharomyces cerevisiae. We are characterizing these mutants to determine the effect these mutations have on Dib1 protein structure and their overall impact on splicing activity. This study will identify the role of critical residues in Dib1 in pre‐mRNA splicing.