Quantitative Analysis of Viral mRNA Translation

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Plant viral diseases affect a significant number of food crops world-wide and can have severe impact both on economic conditions and food supply. Protein synthesis is a key step in viral infection yet is not well understood. This research will increase our understanding of plant viral protein synthesis and how this important biochemical process can be regulated. Structural information about IRES and initiation factors is providing information about the complexes formed. However, quantitative data are necessary to determine how initiation factors compete for different mRNA sites, the formation of intermediates, and the kinetic and thermodynamic stability of complexes. This research will utilize fluorescence spectroscopy, stopped-flow kinetics and circular dichroism to determine the role of individual initiation factors in plant viral mRNA internal initiation, the interaction of these factors with each other and known IRES elements, and how these factors modify mRNA structure. These data will be used to determine a model for selection of mRNA under various physiologic conditions. Quantitative information and determination of the cooperativity of interactions will yield detailed information on the assembly of the 48S initiation complex and binding to viral IRES. This information can potentially be used to shut off viral protein synthesis either through genetic engineering or biochemical agents. Further, viral protein synthesis in general is simpler than host cell protein synthesis so there is potential to use the information from this research to develop systems to produce desired proteins which are nutritionally beneficial, antibodies, or have other economic uses.