Cation-dependent folding of 3' cap-independent translation elements facilitates interaction of a 17-nucleotide conserved sequence with eIF4G.
The 3'-untranslated regions of many plant viral RNAs contain cap-independent translation elements (CITEs) that drive translation initiation at the 5'-end of the mRNA. The barley yellow dwarf virus-like CITE (BTE) stimulates translation by binding the eIF4G subunit of translation initiation factor eIF4F with high affinity. To understand this interaction, we characterized the dynamic structural properties of the BTE, mapped the eIF4G-binding sites on the BTE and identified a region of eIF4G that is crucial for BTE binding. BTE folding involves cooperative uptake of magnesium ions and is driven primarily by charge neutralization. Footprinting experiments revealed that functional eIF4G fragments protect the highly conserved stem-loop I and a downstream bulge. The BTE forms a functional structure in the absence of protein, and the loop that base pairs the 5'-untranslated region (5'-UTR) remains solvent-accessible at high eIF4G concentrations. The region in eIF4G between the eIF4E-binding site and the MIF4G region is required for BTE binding and translation. The data support the model in which the eIF4F complex binds directly to the BTE which base pairs simultaneously to the 5'-UTR, allowing eIF4F to recruit the 40S ribosomal subunit to the 5'-end.
Department of Plant Pathology and Microbiology, Iowa State University, Ames, IA 50011, USA.
Jelena J
JJ
Kraft
Krzysztof
K
Treder
Mariko S
MS
Peterson
W Allen
WA
Miller
D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D039603:Eukaryotic Initiation Factor-4G / Q000737:chemistry / Q000235:genetics*; D015870:Gene Expression; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter; D049409:Luciferases, Firefly / Q000096:biosynthesis / Q000235:genetics; D017793:Luteovirus / Q000235:genetics*; D015636:Magnesium Chloride / Q000737:chemistry; D008969:Molecular Sequence Data; D010442:Peptide Chain Initiation, Translational; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics*; D011188:Potassium / Q000737:chemistry; D011485:Protein Binding; D059370:RNA Folding; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics*; D038621:Regulatory Sequences, Ribonucleic Acid; D020506:Untranslated Regions
D039603:Eukaryotic Initiation Factor-4G; D010940:Plant Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D038621:Regulatory Sequences, Ribonucleic Acid; D020506:Untranslated Regions; D015636:Magnesium Chloride; D049409:Luciferases, Firefly; D011188:Potassium