Document Detail

Structural elements in the 5'-untranslated region of giardiavirus transcript essential for internal ribosome entry site-mediated translation initiation.
MedLine Citation:
PMID:  15821134     Owner:  NLM     Status:  MEDLINE    
Translation of uncapped giardiavirus (GLV) mRNA in Giardia lamblia requires the presence of a 5'-untranslated region (5'-UTR) and a viral capsid coding region. We used dicistronic viral constructs to show that the downstream 253 nucleotides (nt) of the 5'-UTR plus the initial 264-nt capsid coding region constitute an internal ribosome entry site (IRES). Predicted secondary structures in the 253-nt 5'-UTR include stem-loops U3, U4a, U4b, U4c, and U5. Chemical and enzymatic probing analysis confirmed the presence of all predicted stem-loops except U4a. Disruption of stem-loop structures U3 and U5 by site-directed mutagenesis resulted in a drastic reduction in translation of a monocistronic viral transcript, which could be restored by compensatory sequence changes. Mutations disrupting stem-loops U4b and U4c do not exert an appreciable effect on translation, but certain sequences in the U4a region and in U4b do appear to play important roles in the IRES. Structural analysis also suggests that an 8-nt U3 loop sequence (nt 147 to 154) pairs with an 8-nt downstream sequence (nt 168 to 175) to form a pseudoknot. Disruption of this pseudoknot by mutagenesis resulted in a drastic reduction in translation, which could be restored by compensatory sequence changes. This study has defined the secondary structure in the 5'-UTR of the IRES. Together with the previous results, we have now completed analysis of the entire structure of GLV IRES and fully defined the functionally essential structural elements in it.
Srinivas Garlapati; Ching C Wang
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Eukaryotic cell     Volume:  4     ISSN:  1535-9778     ISO Abbreviation:  Eukaryotic Cell     Publication Date:  2005 Apr 
Date Detail:
Created Date:  2005-04-11     Completed Date:  2005-08-30     Revised Date:  2013-06-09    
Medline Journal Info:
Nlm Unique ID:  101130731     Medline TA:  Eukaryot Cell     Country:  United States    
Other Details:
Languages:  eng     Pagination:  742-54     Citation Subset:  IM    
Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94107-2280, USA.
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MeSH Terms
5' Untranslated Regions / genetics*
Base Pairing
Base Sequence
Capsid / physiology*
Codon, Initiator / genetics
Giardia lamblia / genetics,  virology*
Giardiavirus / genetics*
Molecular Sequence Data
Mutagenesis, Site-Directed
Nucleic Acid Conformation*
Peptide Chain Initiation, Translational
Protein Biosynthesis / genetics*
RNA Stability
RNA, Messenger / chemistry,  genetics,  metabolism
RNA, Viral / chemistry*,  genetics,  metabolism
Ribosomes / genetics
Structure-Activity Relationship
Grant Support
Reg. No./Substance:
0/5' Untranslated Regions; 0/Codon, Initiator; 0/RNA, Messenger; 0/RNA, Viral

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