Domains on the hepatitis C virus internal ribosome entry site for 40s subunit binding.
The internal ribosome entry site (IRES) of the hepatitis C virus (HCV) RNA is known to interact with the 40S ribosomal subunit alone, in the absence of any additional initiation factors or Met-tRNAi. Previous work from this laboratory on the 80S and 48S ribosomal initiation complexes involving the HCV IRES showed that stem-loop III, the pseudoknot domain, and some coding sequence were protected from pancreatic RNase digestion. Stem-loop II is never protected by these complexes. Furthermore, there is no prior evidence reported showing extensive direct binding of stem-loop II to ribosomes or subunits. Using direct analysis of RNase-protected HCV IRES domains bound to 40S ribosomal subunits, we have determined that stem-loops II and III and the pseudoknot of the HCV IRES are involved in this initial binding step. The start AUG codon is only minimally protected. The HCV-40S subunit binary complex thus involves recognition and binding of stem-loop II, revealing its role in the first step of a multistep initiation process that may also involve rearrangement of the bound IRES RNA as it progresses.
Lytle
J Robin
JR
Department of Biochemistry, Weill Medical College of Cornell University, New York, New York 10021, USA.
Wu
Lily
L
Robertson
Hugh D
HD
D020121:5' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D066298:In Vitro Techniques; D008954:Models, Biological; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D014176:Protein Biosynthesis; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011817:Rabbits; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism
D020121:5' Untranslated Regions; D012367:RNA, Viral