title: An inhibitor of eIF2 activity in the sRNA pool of eukaryotic cells.
abstract: Eukaryotic protein synthesis is a multi-step and highly controlled process that includes an early initiation complex containing eukaryotic initiation factor 2 (eIF2), GTP, and methionine-charged initiator methionyl-tRNA (met-tRNAi). During studies to reconstruct formation of the ternary complex containing these molecules, we detected a potent inhibitor in low molecular mass RNA (sRNA) preparations of eukaryotic tRNA. The ternary complex inhibitor (TCI) was retained in the total sRNA pool after met-tRNAi was charged by aminoacyl tRNA synthetase, co-eluted with sRNA by size exclusion chromatography, but resolved from met-tRNAi by ion exchange chromatography. The adverse effect of TCI was not overcome by high GTP or magnesium omission and was independent of GTP regeneration. Rather, TCI suppressed the rate of ternary complex formation, and disrupted protein synthesis and the accumulation of heavy polymeric ribosomes in reticulocyte lysates in vitro. Lastly, a component or components in ribosome depleted cell lysate significantly reversed TCI activity. Since assembly of the met-tRNAi/eIF2/GTP ternary complex is integral to protein synthesis, awareness of TCI is important to avoid confusion in studies of translation initiation. A clear definition of TCI may also allow a better appreciation of physiologic or pathologic situations, factors, and events that control protein synthesis in vivo.
authors.affiliation: Department of Surgery, Yale University School of Medicine, New Haven, CT 06520-8041, USA. michael.centrella@yale.edu
authors.forename: Michael
authors.initials: M
authors.lastname: Centrella
authors.forename: David L
authors.initials: DL
authors.lastname: Porter
authors.forename: Thomas L
authors.initials: TL
authors.lastname: McCarthy
mesh_terms: D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D000818:Animals; D005057:Eukaryotic Cells / Q000187:drug effects / Q000378:metabolism*; D015852:Eukaryotic Initiation Factor-2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006868:Hydrolysis / Q000187:drug effects; D008274:Magnesium / Q000494:pharmacology; D051379:Mice; D011132:Polyribosomes / Q000187:drug effects / Q000378:metabolism; D014176:Protein Biosynthesis / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D012313:RNA / Q000378:metabolism*; D012343:RNA, Transfer / Q000378:metabolism; D011817:Rabbits
chemical_list: D015852:Eukaryotic Initiation Factor-2; D012313:RNA; D006160:Guanosine Triphosphate; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases; D008274:Magnesium