The Role of Eukaryotic Initiation Factor (eIF) 4G in Initiation of Translation

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Hellen MCB 9726958 1. Technical Eukaryotic initiation factor (eIF) 4G coordinates the rate-limiting step in protein synthesis by interacting with other factors in the recruitment of mRNA to ribosomes. eIF4G associates with the cap-binding protein eIF4E and the ATPase/RNA helicase eIF4A to form the eIF4F complex. eIF4G also binds to eIF3, a component of the 43S ribosomal preinitiation complex. This study applies biochemical and molecular genetic approaches to characterize the role of eIF4G' in initiation of translation. This is achieved through reconstitution experiments in vitro using purified components, which include recombinant eIFs 4A, 4B, 4E and 4G, on a variety of clone mRNAs. Sensitive toe-printing and footprinting methods to map the position of ribosomal and preinitiation complexes on mRNAs are developed for this purpose. The significance of this study lies in its advancing our knowledge of protein synthesis at the molecular level. 2. Non Technical This study is to elucidate fundamental aspects of the mechanism and regulation of protein synthesis, particularly the role of the initiation factor, 4G (eIF4G), in coordinating the activities of various cellular components that together recruit individual mRNAs for the onset of translation. Experimentally, purified components on different mRNAs are reconstituted in vitro at the beginning stage of translation. Various protein or mRNA components are systematically substituted with a series of variants, and their effects compared under specific conditions such as growth arrest and mitosis. The importance of determinants in eIF4G that enable it to bind to all mRNAs nonspecifically, and to a few with high specificity, are identified and characterized. Viral and cellular mRNAs that bind eIF4G with high affinity are chosen in this study, since they may be models for translation when eIF4E is inactive. Also being studied are determinants of eIF4A's which interact with eIF4G, another initiation factor. This interaction directs the unwinding of eIF4A to a specific site on the mRNA to permit the attachment of ribosomes, the anchorage machinery for protein synthesis.