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Ribosomal release without peptidyl tRNA hydrolysis at translation termination in a eukaryotic system

Published online by Cambridge University Press:  01 February 1998

JIANHONG CAO  and
ADAM P. GEBALLE
JIANHONG CAO
Affiliation:
Divisions of Molecular Medicine and Clinical Research, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA
ADAM P. GEBALLE
Affiliation:
Divisions of Molecular Medicine and Clinical Research, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA Department of Medicine and Department of Microbiology, University of Washington, Seattle, Washington 98195, USA
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Abstract

A 22-codon upstream open reading frame (uORF2) in the human cytomegalovirus UL4 transcript leader inhibits downstream translation in cis. Previous studies revealed that the peptide product of uORF2 mediates this inhibitory effect by interfering with translation termination at its own stop codon. The block at termination results both in accumulation of the nascent uORF2 peptide linked to tRNAPro, the tRNA that decodes the final codon of uORF2, and in stalling of ribosomes at the end of uORF2. The stalled ribosomes create a barrier that obstructs ribosomal transit to the downstream cistron. In the current studies, we further investigated the mechanism of uORF2-mediated translational inhibition by assessing the kinetics of uORF2 peptidyl tRNAPro hydrolysis and ribosomal release from the uORF2 termination site. Whereas hydrolysis of a mutant, noninhibitory uORF2 peptidyl tRNA is nearly complete in less than 1 min, hydrolysis of the wild-type peptidyl tRNA is negligible even after 30 min. In spite of this remarkably prolonged block to hydrolysis of the uORF2 peptidyl tRNAPro, most ribosomes are released from the uORF2 termination site within 15 min. Thus, peptidyl tRNA hydrolysis is not absolutely required for ribosomal release in this system. These results suggest that a eukaryotic cellular mechanism exists for removing stalled ribosomes from mRNAs in the absence of peptidyl tRNA hydrolysis.

Keywords

cytomegalovirustoeprinttranslational controlUL4upstream open reading frame
Type
Research Article
Information
RNA , Volume 4 , Issue 2 , February 1998 , pp. 181 - 188
Copyright
© 1998 RNA Society

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