Transcriptional organization and regulation of the Escherichia coli K30 group 1 capsule biosynthesis (cps) gene cluster

Abstract

SummaryEscherichia coli group 1 capsules are important virulence determinants, yet little is known about the transcriptional organization or regulation of their biosynthetic ( cps ) operons. Transcription of the prototype serotype K30 cluster is modulated by the JUMPStart-RfaH antitermination mechanism, with the cps promoter being localized to a region immediately upstream of the JUMPStart sequence. A putative stem-loop structure located within the K30 cps cluster separates conserved genes with products that are required for surface expression of capsule from serotype-specific genes encoding enzymes for polymer repeat-unit synthesis and polymerization. This putative stem-loop structure significantly reduces transcription in a termination-probe vector and may contribute to differential expression of the cps genes. Previous work indicated that increased amounts of group 1 capsular polysaccharide synthesis resulted from the overexpression of the Rcs (regulator of capsule synthesis) proteins. However, neither overexpression of the transcriptional activator RcsB nor an rcsB :: aadA chromosomal insertion altered the level of transcription measured by cps::lacZ fusions. In the group 1 strains examined, an RcsAB box was found immediately upstream of galF , a gene involved in the production of sugar nucleotide precursors. Overexpression of RcsB was found to result in a threefold increase in transcription of a galF::lacZ chromosomal fusion. Moreover, overexpression of GalF gave rise to a two-to threefold increase in cell-free as well as cellassociated capsule, without affecting cps::lacZ activity. These results indicate that transcription of the E. coli group 1 capsule cluster itself is not regulated by the Rcs system and may, in fact, be constitutive. However, the Rcs system can potentially influence levels of capsular polysaccharide production by increasing galF transcription and influencing the available pool of biosynthetic precursors.

The UDP-glucose Dehydrogenase of Escherichia coli K-12 Displays Substrate Inhibition by NAD That Is Relieved by Nucleotide Triphosphates

“…In contrast, group 1 CPSs are constitutively expressed in E. coli and K. pneumoniae, where they are important virulence factors. The cps locus is not responsive to RcsC/YojN/RcsB/(RcsA) (23) in E. coli with group 1 CPSs. Enhanced CPS production in these strains results from Rcs-mediated up-regulation of the galF gene, whose product is implicated in the biosynthesis of a UDPglucose precursor (55).…”

“…Colanic acid is an environmentally regulated exopolysaccharide, which is activated by the Rcs regulon (22). Group 1 CPSs are constitutively expressed virulence determinants (23) and are encoded by gene clusters that occupy the same chromosomal location as the colanic acid genes (24). Consequently, possession of colanic acid and group 1 capsule assembly machinery appears to be mutually exclusive.…”

Regulation of bacterial virulence gene expression by cell envelope stress responses

“…Alternatively, perhaps the appropriate environmental conditions were not used to detect an effect of the rcsB null mutation. 115 Finally, an analogous situation also occurs in Salmonella. In this case, the ugd gene, encoding an enzyme that converts UDP-dglucose to UDP-d-glucuronic acid, is regulated by the Rcs system.…”

“…115 Independent overexpression of galF also increased capsule production without affecting expression of the cps operon. 115 Thus, the Rcs system might be able to induce group I capsule by increasing galF expression and influencing the pool of precursors. This does not explain why an rcsB null mutation did not affect capsule production.…”

“…Indeed, it was found that rcsB overexpression induced the galF gene involved in producing sugar nucleotide precursors, and that there was a putative RcsAB-binding sequence (RcsAB box) upstream of galF. 115 Independent overexpression of galF also increased capsule production without affecting expression of the cps operon. 115 Thus, the Rcs system might be able to induce group I capsule by increasing galF expression and influencing the pool of precursors.…”