Ribosome Recycling Factor and Release Factor 3 Action Promotes TnaC-Peptidyl-tRNA Dropoff and Relieves Ribosome Stalling during Tryptophan Induction of
tna
Operon Expression in
Escherichia coli
Authors: Gong Ming , Cruz-Vera Luis R. , Yanofsky Charles 
Published: February 2007

ABSTRACT

Upon tryptophan induction of
tna
operon expression in
Escherichia coli
, the leader peptidyl-tRNA, TnaC-

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, resists cleavage, resulting in ribosome stalling at the
tnaC
stop codon. This stalled ribosome blocks Rho factor binding and action, preventing transcription termination in the
tna
operon's leader region. Plasmid-mediated overexpression of
tnaC
was previously shown to inhibit cell growth by reducing uncharged

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availability. Which factors relieve ribosome stalling, facilitate TnaC-

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cleavage, and relieve growth inhibition were addressed in the current study. In strains containing the chromosomal
tna
operon and lacking a
tnaC
plasmid, the overproduction of ribosome recycling factor (RRF) and release factor 3 (RF3) reduced
tna
operon expression. Their overproduction in vivo also increased the rate of cleavage of TnaC-

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, relieving the growth inhibition associated with plasmid-mediated
tnaC
overexpression. The overproduction of elongation factor G or initiation factor 3 did not have comparable effects, and tmRNA was incapable of attacking TnaC-

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in stalled ribosome complexes. The stability of TnaC-

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was increased appreciably in strains deficient in RRF and RF3 or deficient in peptidyl-tRNA hydrolase. These findings reveal the existence of a natural mechanism whereby an amino acid, tryptophan, binds to ribosomes that have just completed the synthesis of TnaC-

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. Bound tryptophan inhibits RF2-mediated cleavage of TnaC-

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, resulting in the stalling of the ribosome translating
tnaC
mRNA. This stalling results in increased transcription of the structural genes of the
tna
operon. RRF and RF3 then bind to this stalled ribosome complex and slowly release TnaC-

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. This release allows ribosome recycling and permits the cleavage of TnaC-

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by peptidyl-tRNA hydrolase.