The regulation of gene expression is critical for cell growth and development. The success of transcriptional and post-transcriptional mechanisms that regulate gene expression hinge upon the successful synthesis of functional polypeptides. Many soluble translation factors, the aminoacyltRNAs (aa-tRNAs) and the ribosome interact to produce nascent polypeptides chains required by the cell. This proposal uses the translation Elongation Factor eEFIA as a central point from which to examine the interactions between these critical components of the translation machinery that function to ensure accurate and appropriate protein synthesis. The analysis of eEF1A function will dissect the mechanisms by which translational fidelity is ensured and how eEF1A serves to integrate the communication between the aa-tRNA and the ribosome. As a G-protein, work on eEF1A has implications in our understanding of cellular regulatory mechanism through nucleotide exchange and hydrolysis. The yeast Saccharomyces cerevisiae allows a combined genetic and biochemical approach to address key questions regarding the mechanism of elongation and the maintenance of translational accuracy. While these studies have implications for all eukaryotic systems, the interaction of eEF1A with the fungal specific translation factor eEF3 defines a process unique to fungi. The interaction of eEF3 with eEF1A and the ribosome, and its essential function in vivo, support the role of eEF3 as a key player in fungal protein synthesis. Roles for eEF3 in translational accuracy have also been proposed, however, little is known about the effects of eEF3 on fidelity in vivo. Accurate protein synthesis is a critical step in ensuring the regulatory mechanisms that exist for modulating gene expression are productive. Interestingly, the accuracy of translation elongation is a process many viruses manipulate to complete a critical part of their life cycle. The proposed studies address the critical communication involved in ensuring translational accuracy in eukaryotes, while potentially yielding new insights into viral-host interactions. Additionally, the analysis of a fungal specific translation factor enhances our understanding of a unique aspect of protein synthesis and may yield new anti-fungal targets.
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