Functional Insights Into the Role of gppA in (p)ppGpp Metabolism of Vibrio cholerae

Abstract

The stringent response, an adaptive response to nutrient limitation and exposure to xenobiotics in bacteria, is mediated by two intracellular signaling molecules, pppGpp and ppGpp, together called (p)ppGpp. The cellular level of (p)ppGpp in bacterial cells is controlled by the Rel/Spo family of proteins. In the cholera pathogen, Vibrio cholerae, (p)ppGpp metabolism is regulated by the products of at least three genes relA, spoT, and relV. In this study, we identify and characterize the function of the guanosine-5-triphosphate 3-diphosphate pyrophosphatase A (GppA) encoding gene gppA of V. cholerae. Genomic analysis indicates that the gppA locus is conserved in vibrios and organized as a bicistronic operon along with the rhlB gene. We engineered the genome of V. cholerae to develop different mutants devoid of GppA and/or other phosphate metabolic enzymes. Our findings indicate that in V. cholerae, GppA plays an important role in the conversion of pppGpp to ppGpp during amino acid deprivation but not during glucose starvation. Quantitative analyses of the gppA transcript level reveal its differential expression pattern at different growth phases and starvation conditions. It has been observed that the GppA deficiency during amino acid starvation condition could be complemented by overexpressing the exopolyphosphatase coding gene ppx of V. cholerae. By deletion analysis, we further demonstrate that the amino and carboxy terminal sequences flanking the Ppx-GppA motif of the GppA protein of V. cholerae are also important for its enzymatic function.

Proteome Expression and Survival Strategies of a Proteorhodopsin-ContainingVibrioStrain under Carbon and Nitrogen Limitation

“…However, both media had the same replete concentrations of phosphate, suggesting that these observed expression patterns may in fact be responses to carbon stress, mediated by the stringent response, which is recognized to be involved in dormancy transitions ( 26 ). The stringent stress response in many bacteria, including Vibrio , is signaled by intracellular accumulation of (p)ppGpp synthesized by RelA and/or SpoT ( 55 ), which modulates the stability and activity of a wide array of transcription factors, including RpoS. In E. coli ’s stringent response, induction of phoA and pstS in response to phosphate starvation are dependent on ppGpp synthesized via the spoT pathway, not relA ( 56 , 57 ).…”

Whole-genome sequencing analysis of two heat-evolved Escherichia coli strains

“…ppGpp and pppGpp especially inhibit polyphosphate degradation by the exopolyphosphatase Ppx, and Gpp also converts pppGpp to ppGpp. The gpp frame shift is far enough into the gene that the encoded protein retains the domains required for conversion of pppGpp to ppGpp and for polyphosphatase activity (frame shifted after amino acid 387 of 494) [ 68 , 69 ]. However, the frameshift occurs partway through one of the C-terminal domains, indicating that this frameshift could cause abnormal folding and therefore perhaps degradation of Gpp.…”