Regulatory Effects of CsrA in Vibrio cholerae

Abstract

CsrA is a posttranscriptional global regulator in Vibrio cholerae. Although CsrA is critical for V. cholerae survival within the mammalian host, the regulatory targets of CsrA remain mostly unknown. To identify pathways controlled by CsrA, RNA-seq transcriptome analysis was carried out by comparing the wild type and the csrA mutant grown to early exponential, mid-exponential, and stationary phases of growth. This enabled us to identify the global effects of CsrA-mediated regulation throughout the V. cholerae growth cycle. We found that CsrA regulates 22% of the V. cholerae transcriptome, with significant regulation within the gene ontology (GO) processes that involve amino acid transport and metabolism, central carbon metabolism, lipid metabolism, iron uptake, and flagellum-dependent motility. Through CsrA-RNA coimmunoprecipitation experiments, we found that CsrA binds to multiple mRNAs that encode regulatory proteins. These include transcripts encoding the major sigma factors RpoS and RpoE, which may explain how CsrA regulation affects such a large proportion of the V. cholerae transcriptome. Other direct targets include flrC, encoding a central regulator in flagellar gene expression, and aphA, encoding the virulence gene transcription factor AphA. We found that CsrA binds to the aphA mRNA both in vivo and in vitro, and CsrA significantly increases AphA protein synthesis. The increase in AphA was due to increased translation, not transcription, in the presence of CsrA, consistent with CsrA binding to the aphA transcript and enhancing its translation. CsrA is required for the virulence of V. cholerae and this study illustrates the central role of CsrA in virulence gene regulation. IMPORTANCE Vibrio cholerae, a Gram-negative bacterium, is a natural inhabitant of the aqueous environment. However, once ingested, this bacterium can colonize the human host and cause the disease cholera. In order to successfully transition between its aqueous habitat and the human host, the bacterium must sense changes in its environment and rapidly alter gene expression. Global regulators, including CsrA, play an integral role in altering the expression of a large number of genes to promote adaptation and survival, which is required for intestinal colonization. We used transcriptomics and a directed CsrA-RNA coimmunoprecipitation to characterize the CsrA regulon and found that CsrA alters the expression of more than 800 transcripts in V. cholerae. Processes regulated by CsrA include motility, the rugose phenotype, and virulence pathways. CsrA directly binds to the aphA transcript and positively regulates the production of the virulence regulator AphA. Thus, CsrA regulates multiple processes that have been linked to pathogenesis.

The Escherichia coli two-component signal sensor BarA binds protonated acetate via a conserved hydrophobic-binding pocket

“…This signaling circuitry is highly conserved in γ-proteobacteria ( 42 , 43 , 44 ), preserving most of its general characteristics. However, the genes whose expression are modulated by the BarA/UvrY/CsrA regulatory cascade and hence the physiological response, may vary significantly among bacteria ( 10 , 45 , 46 , 47 , 48 , 49 , 50 , 51 ). Here, we found that the input domain of the BarA/UvrY system of E. coli differs from that of the homologous GacS/GacA of P. aeruginosa, in that the former has evolved to recognize an uncharged signal.…”

Specific and Global RNA Regulators in Pseudomonas aeruginosa

“…In fact, the repressive function of Hfq can be antagonized through sequestration by the regulatory RNA CrcZ, containing multiple A-rich motifs, which in turn results in de-repression of Hfq and Hfq/Crc target genes (Figures 2 and 3b). The CsrA/RsmA family of RNA-binding proteins governs virulence gene expression in pathogenic bacteria [173][174][175][176][177][178], including Pae [139,179]. Members of this family usually negatively regulate translation of target mRNAs through binding to stem-loop structures present in their 5 UTR that contain exposed GGA motifs [27,139,173,[180][181][182][183][184].…”

“…The CsrA/RsmA family of RNA-binding proteins governs virulence gene expres in pathogenic bacteria [173][174][175][176][177][178], including Pae [139,179]. Members of this family usu negatively regulate translation of target mRNAs through binding to stem-loop struct present in their 5′UTR that contain exposed GGA motifs [27,139,173,[180][181][182][183][184].…”

The VarA-CsrA regulatory pathway influences cell shape in Vibrio cholerae

“…The VarS/VarA system also contributes to the dissemination of V. cholerae from the host into the environment (10). Moreover, in conjunction with CsrA, this TCS is known to be involved in the regulation of central carbon metabolism, iron uptake, lipid metabolism, flagellum-dependent motility, and other phenotypes (11,12). Collectively, signaling through the VarS/VarA-CsrA circuit therefore affects the environmental lifestyle of V. cholerae as well as its pathogenesis.…”