title: Cryptic AUG is important for 48S ribosomal assembly during internal initiation of translation of coxsackievirus B3 RNA.
abstract: We have investigated the possible role of a conserved cis-acting element, the cryptic AUG, present in the 5' UTR of coxsackievirus B3 (CVB3 ) RNA. CVB3 5' UTR contains multiple AUG codons upstream of the initiator AUG, which are not used for the initiation of translation. The 48S ribosomal assembly takes place upstream of the cryptic AUG. We show here that mutation in the cryptic AUG results in reduced efficiency of translation mediated by the CVB3 IRES; mutation also reduces the interaction of mutant IRES with a well characterized IRES trans-acting factor, the human La protein. Furthermore, partial silencing of the La gene showed a decrease in IRES activity in the case of both the wild-type and mutant. We have demonstrated here that the interaction of the 48S ribosomal complex with mutant RNA was weaker compared with wild-type RNA by ribosome assembly analysis. We have also investigated by chemical and enzymic modifications the possible alteration in secondary structure in the mutant RNA. Results suggest that the secondary structure of mutant RNA was only marginally altered. Additionally, we have demonstrated by generating compensatory and non-specific mutations the specific function of the cryptic AUG in internal initiation. Results suggest that the effect of the cryptic AUG is specific and translation could not be rescued. However, a possibility of tertiary interaction of the cryptic AUG with other cis-acting elements cannot be ruled out. Taken together, it appears that the integrity of the cryptic AUG is important for efficient translation initiation by the CVB3 IRES RNA.
authors: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors: Bhupendra
authors: B
authors: Verma
authors: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors: Anand
authors: A
authors: Ponnuswamy
authors: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors: Sivakumar Vadivel
authors: SV
authors: Gnanasundram
authors: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors: Saumitra
authors: S
authors: Das
mesh_terms: D020121:5' Untranslated Regions; D018387:Codon, Initiator; D029822:Enterovirus B, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D010442:Peptide Chain Initiation, Translational; D012367:RNA, Viral / Q000235:genetics*; D014764:Viral Proteins / Q000096:biosynthesis*
chemical_list: D020121:5' Untranslated Regions; D018387:Codon, Initiator; D012367:RNA, Viral; D014764:Viral Proteins