title: The Triticum Mosaic Virus 5' Leader Binds to Both eIF4G and eIFiso4G for Translation.
abstract: We recently identified a remarkably strong (739 nt-long) IRES-like element in the 5' untranslated region (UTR) of Triticum mosaic virus (TriMV, Potyviridae). Here, we define the components of the cap-binding translation initiation complex that are required for TriMV translation. Using bio-layer interferometry and affinity capture of the native translation apparatus, we reveal that the viral translation element has a ten-fold greater affinity for the large subunit eIF4G/eIFiso4G than to the cap binding protein eIF4E/eIFiso4E. This data supports a translation mechanism that is largely dependent on eIF4G and its isoform. The binding of both scaffold isoforms requires an eight base-pair-long hairpin structure located 270 nucleotides upstream of the translation initiation site, which we have previously shown to be crucial for IRES activity. Despite a weak binding affinity to the mRNA, eIFiso4G alone or in combination with eIFiso4E supports TriMV translation in a cap-binding factor-depleted wheat germ extract. Notably, TriMV 5' UTR-mediated translation is dependent upon eIF4A helicase activity, as the addition of the eIF4A inhibitor hippuristanol inhibits 5' UTR-mediated translation. This inhibition is reversible with the addition of recombinant wheat eIF4A. These results and previous observations demonstrate a key role of eIF4G and eIF4A in this unique mechanism of cap-independent-translation. This work provides new insights into the lesser studied translation mechanisms of plant virus-mediated internal translation initiation.
authors.affiliation: Department of Plant Pathology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors.forename: Robyn
authors.initials: R
authors.lastname: Roberts
authors.affiliation: Department of Molecular Biosciences, University of Texas Austin, Austin, Texas, United States of America.
authors.forename: Laura K
authors.initials: LK
authors.lastname: Mayberry
authors.affiliation: Department of Plant Pathology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
authors.forename: Karen S
authors.initials: KS
authors.lastname: Browning
authors.affiliation: Department of Molecular Biosciences, University of Texas Austin, Austin, Texas, United States of America.
authors.forename: Aurélie M
authors.initials: AM
authors.lastname: Rakotondrafara
mesh_terms: D020121:5' Untranslated Regions; D039603:Eukaryotic Initiation Factor-4G / Q000737:chemistry*; D006801:Humans; D007368:Interferometry; D009029:Mosaic Viruses / Q000235:genetics; D019176:Potyviridae / Q000235:genetics*; D011485:Protein Binding; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000737:chemistry; D012315:RNA Caps / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry; D012270:Ribosomes / Q000737:chemistry; D013261:Sterols / Q000737:chemistry; D014908:Triticum / Q000821:virology*; D055544:beta-Globins / Q000737:chemistry
chemical_list: D020121:5' Untranslated Regions; D039603:Eukaryotic Initiation Factor-4G; D020033:Protein Isoforms; D012315:RNA Caps; D012333:RNA, Messenger; D012367:RNA, Viral; D011994:Recombinant Proteins; D013261:Sterols; D055544:beta-Globins; C510125:hippuristanol