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J Biol Chem, Vol. 273, Issue 9, 5006-5012, February 27, 1998
From the Department of Biochemistry and Molecular Biology,
Louisiana State University Medical Center,
Shreveport, Louisiana 71130-3932
The eIF4 group initiation factors are required
for cap-dependent translation initiation. Infection of
mammalian cells by picornaviruses results in proteolytic cleavage of
one of these factors, eIF4G, which severely restricts
cap-dependent initiation but permits cap-independent
initiation to proceed from an internal ribosome entry site (IRES) in
picornaviral RNAs. The first 357 nucleotides (nt) of the
5'-untranslated region of eIF4G mRNA also contains an IRES. Using
bicistronic constructs for expression in K562 cells, we have now shown
that progressive deletions of the 5'-untranslated region can have
either stimulatory or inhibitory effects. Furthermore, a 101-nt segment
exhibits full IRES activity, and an 81-nt segment exhibits detectable
IRES activity. A polypyrimidine tract (PPT) at the 3' terminus is
essential for internal initiation, a property which is characteristic
of picornaviral IRESs but not the other host cellular IRESs studied to
date. IRES activity does not require sequences beyond 357 nt.
Out-of-frame AUGs have no effect on IRES-driven luciferase expression
when introduced upstream of the PPT but markedly decrease expression
when introduced at sites between the PPT and the authentic initiation
codon at nt 369. These results suggest that the ribosomal subunit
enters at or near the PPT and then scans downstream for the initiation
codon.
Functional Characterization of the Internal Ribosome Entry Site
of eIF4G mRNA
Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
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