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Mol Microbiol. 2008 Sep;69(6):1439-49. doi: 10.1111/j.1365-2958.2008.06370.x. Epub 2008 Jul 18.

Erythromycin-induced ribosome stalling and RNase J1-mediated mRNA processing in Bacillus subtilis.

Author information

1
Department of Pharmacology and Systems Therapeutics, Mount Sinai School of Medicine of New York University, New York, NY 10029, USA.

Abstract

Addition of erythromycin (Em) to a Bacillus subtilis strain carrying the ermC gene results in ribosome stalling in the ermC leader peptide coding sequence. Using DeltaermC, a deletion derivative of ermC that specifies the 254 nucleotide DeltaermC mRNA, we showed previously that ribosome stalling is concomitant with processing of DeltaermC mRNA, generating a 209 nucleotide RNA whose 5' end maps to codon 5 of the DeltaermC coding sequence. Here we probed for peptidyl-tRNA to show that ribosome stalling occurs after incorporation of the amino acid specified by codon 9. Thus, cleavage upstream of codon 5 is not an example of 'A-site cleavage' that has been reported for Escherichia coli. Analysis of DeltaermC mRNA processing in endoribonuclease mutant strains showed that this processing is RNase J1-dependent. DeltaermC mRNA processing was inhibited by the presence of stable secondary structure at the 5' end, demonstrating 5'-end dependence, and was shown to be a result of RNase J1 endonuclease activity, rather than 5'-to-3' exonuclease activity. Examination of processing in derivatives of DeltaermC that had codons inserted upstream of the ribosome stalling site revealed that Em-induced ribosome stalling can occur considerably further from the start codon than would be expected based on previous studies.

PMID:
18647167
PMCID:
PMC2575084
DOI:
10.1111/j.1365-2958.2008.06370.x
[Indexed for MEDLINE]
Free PMC Article

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