Vitamin B₁₂ repression of the cob operon in Salmonella typhimurium: translational control of the cbiA gene

Abstract

Expression of the cob operon is repressed by B12 via a post-transcriptional control mechanism which requires sequence elements within the leader region of the mRNA and the first gene of the operon, the cbiA gene. Here we show that B12 repression of cbiA gene expression occurs at the level of translation initiation through sequestration of the ribosomal binding site (rbs) in an RNA hairpin. Analysis of mutations that destabilize or restabilize the secondary structure demonstrates that folding of the hairpin is essential for repression. The existence of the hairpin was confirmed by a secondary structure analysis of RNA from the wild type and three mutants. Deletions that remove the upstream part of the leader confer a drastic reduction in translation efficiency. This low-level translation is caused by the hairpin, as indicated by the finding that suppressor mutations that destabilize the hairpin restore efficient translation. Thus, the native upstream RNA functions as a translation enhancer and acts to relieve the hairpin's inhibitory effect on translation initiation. The inhibitory effect of the hairpin was confirmed by a ribosomal toeprinting analysis. We propose that the translational control of the cbiA gene mediates repression of the entire cob operon.

A conserved RNA structure ( thi box) is involved in regulation of thiamin biosynthetic gene expression in bacteria

“…These genes are transcribed with unusually long leaders of 241 nt (btuB) or 468 nt (cob operon). Several mutations that led to decreased repression have been mapped to these leaders (40)(41)(42). The B 12 box, a 25-nt element that is conserved in the leaders of the btuB and cbiA genes, is required for the expression of btuB (41,43), as the thi box is required for thiamin-regulated expression of thi genes.…”

Posttranscriptional Regulation of Mammalian Methionine Synthase by B12

“…In the latter organism, expression of the cob operon involved in the de novo biosynthesis of B,, is repressed (26)(27)(28). In both E. coli and in S. typhimurium, B,, represses the btuB gene product encoding an outer membrane B12 transport protein (29,30).…”

“…In addition, a second common element observed in these transcripts is the presence of sequences in the leader with the potential to form an RNA hairpin which sequesters the RNA binding site. In the presence of B,, an RNA hairpin forms with inhibits translation initiation of the cbiA gene in S. typhimurium (28). Since the regulatory elements for translation are distinct in prokaryotes and eukaryotes, it is not surprising that a consensus B,, box is not observed in the human methionine synthase cDNA.…”

A defective mutant of Salmonella enterica Serovar Gallinarum in cobalamin biosynthesis is avirulent in chickens

“…This conclusion is based on the fact that the cob operon, is induced by propanediol and is coregulated with the pdu operon, which are encoding genes for propanediol degradation (11,41). This control pattern suggests that perhaps propanediol could be used as a useful carbon and energy source which is frequently valuable and available under anaerobic conditions, because it is made as a by-product during degradation of common plant sugars (29,53 (40). However, the exact relation is unknown.…”