Bacteriocin Activity of Streptococcus pneumoniae Is Controlled by the Serine Protease HtrA via Posttranscriptional Regulation

Abstract

The blp locus of a type 6A strain of Streptococcus pneumoniae encodes a two-peptide bacteriocin, pneumocin MN, which mediates intraspecies competition during mouse nasopharyngeal colonization. This locus is regulated by a quorum-sensing mechanism consisting of a dedicated two-component regulatory system and a peptide pheromone. Like most clinical isolates, this type 6A strain can be separated into opaque and transparent colony variants, each playing a different role during pneumococcal infection. In this study, we show that the blp locus is differentially regulated at the posttranscriptional level in pneumococcal opacity variants. Transparent and opaque variants produce equivalent amounts of blpMNPO transcript when stimulated with a synthetic pheromone, but transparent variants have no pneumocin MN-mediated inhibitory activity while opaque variants produce large zones of inhibitory activity. The differential regulation in opacity variants is driven by the two-component regulatory system CiaRH via its regulation of the serine protease HtrA. Transparent mutants deficient in CiaH or HtrA show increased pneumocin MN-mediated inhibition. In addition, these mutants demonstrate alterations in their dose response to a synthetic peptide pheromone, suggesting that HtrA activity impacts pneumocin MN production at the level of signaling. This, in addition to its known effects on competence, suggests that HtrA is a pleiotropic regulator whose protease activity affects several important bacterial pathways. The complex regulation of pneumocins may allow the pneumococcus to reserve the secretion of active peptides for situations where the benefit of their inhibitory activity outweighs the cost of their production.

The HtrA Protease of Streptococcus pneumoniae Controls Density-Dependent Stimulation of the Bacteriocin blp Locus via Disruption of Pheromone Secretion

“…We have shown previously that certain naturally occurring phenotypic variants of S. pneumoniae demonstrate growth-phasedependent expression of HtrA (9). Opaque variants have a decrease in HtrA levels at the late exponential phase which is thought to explain their high levels of pneumocin activity compared with those of transparent variants in which HtrA levels are relatively constant.…”

“…Although the specific trigger for Cia activation is unknown, ciaRH and htrA transcripts are induced during competence and thought to play a role in repression of the competence state (21). Similar to their role in competence, we had previously shown that both CiaRH and HtrA are involved in the repression of pneumocin expression (9). Naturally occurring opacity variants that differ only moderately in relative HtrA expression levels demonstrated clear phenotypic differences in pneumocin-mediated inhibition that were dependent on HtrA activity (9).…”

“…Similar to their role in competence, we had previously shown that both CiaRH and HtrA are involved in the repression of pneumocin expression (9). Naturally occurring opacity variants that differ only moderately in relative HtrA expression levels demonstrated clear phenotypic differences in pneumocin-mediated inhibition that were dependent on HtrA activity (9). In transparent variants with higher htrA expression, addition of sBlpC restored the defect in transcriptional activation of the locus but did not restore pneumocinmediated inhibition.…”

“…HtrA is a member of the DegP family of proteases and has been shown to also play a role in regulating pneumococcal competence and survival during high-temperature growth (10)(11)(12). In the 6A background, HtrA had two distinct roles in reducing pneumocin expression: it both repressed pneumocin production by altering the dose-response for the peptide pheromone BlpC and decreased inhibition by reducing the amount of functional pneumocin (9). The most straightforward hypothesis for this observation is that HtrA activity either directly or indirectly results in the inactivation of either the secretion apparatus or the secreted peptides themselves, including both pheromone and structural bacteriocins.…”

“…Previous studies have demonstrated that the expression of pneumocins in the prototypic pneumocin-producing strain, 6A, is repressed by the outer-surface serine protease, HtrA (9). HtrA is a member of the DegP family of proteases and has been shown to also play a role in regulating pneumococcal competence and survival during high-temperature growth (10)(11)(12).…”