Trans-acting factors and cis-acting sequences in posttranscriptional gene expression

Abstract

The control of gene expression is greatly influenced by regulatory events at post-transcriptional steps. In my studies of these post-transcriptional control steps, I have utilized the tools provided by the model organism Saccharomyces cerevisiae, including genetic and molecular tools as well as the complete genome sequence. Based on genetic evidence, I hypothesized that the products of the SKI2, SKI3, and SKI8 genes were involved in a pathway of mRNA degradation that acts in the 3' to 5' direction. I demonstrated that mutations in any of the three genes lead to a stabilization of an mRNA species that degrades 3' to 5'. I further demonstrated that components of a protein complex, the exosome, were also required for 3' to 5 ' mRNA degradation. I went on to demonstrate that mutations that disrupt 3' to 5' mRNA degradation are synthetically lethal with mutations that disrupt another pathway that operates in the 5' to 3' direction. This last observation leads to two conclusions. First, these two mechanisms are likely to be the only major methods of mRNA degradation in Saccharomyces cerevisiae. Second, mRNA degradation is an essential process in this organism. I developed a computational method that uses statistical analysis of oligonucleotide frequencies to identify potential cis-acting elements. Application of my method to a group of genes allows the identification of sequences that may be involved in directing co-regulation. Unlike similar methods, my method accounts for oligonucleotide usage in the genes that are not observed to be co-regulated, ensuring that elements common to all genes will not be erroneously detected. After demonstrating that the method detected several known splicing elements in a group of genes containing introns, I went on to characterize the performance of the method under several conditions designed to simulate 'real world' experiments. Finally, I utilized the method to identify an element in a group of nuclear genes that encode mitochondrial proteins.