The HP0165-HP0166 Two-Component System (ArsRS) Regulates Acid-Induced Expression of HP1186 α-Carbonic Anhydrase in Helicobacter pylori by Activating the pH-Dependent Promoter

Abstract

The periplasmic ␣-carbonic anhydrase of Helicobacter pylori is essential for buffering the periplasm at acidic pH. This enzyme is an integral component of the acid acclimation response that allows this neutralophile to colonize the stomach. Transcription of the HP1186 ␣-carbonic anhydrase gene is upregulated in response to low environmental pH. A binding site for the HP0166 response regulator (ArsR) has been identified in the promoter region of the HP1186 gene. To investigate the mechanism that regulates the expression of HP1186 in response to low pH and the role of the HP0165-HP0166 two-component system (ArsRS) in this acidinducible regulation, Northern blot analysis was performed with RNAs isolated from two different wild-type H. pylori strains (26695 and 43504) and mutants with HP0165 histidine kinase (ArsS) deletions, after exposure to either neutral pH or low pH (pH 4.5). ArsS-dependent upregulation of HP1186 ␣-carbonic anhydrase in response to low pH was found in both strains. Western blot analysis of H. pylori membrane proteins confirmed the regulatory role of ArsS in HP1186 expression in response to low pH. Analysis of the HP1186 promoter region revealed two possible transcription start points (TSP 1 and TSP 2 ) located 43 and 11 bp 5 of the ATG start codon, respectively, suggesting that there are two promoters transcribing the HP1186 gene. Quantitative primer extension analysis showed that the promoter from TSP 1 (43 bp 5 of the ATG start codon) is a pH-dependent promoter and is regulated by ArsRS in combating environmental acidity, whereas the promoter from TSP 2 may be responsible for control of the basal transcription of HP1186 ␣-carbonic anhydrase.

Characterization of a Mesorhizobium loti α-Type Carbonic Anhydrase and Its Role in Symbiotic Nitrogen Fixation

“…The possible role of CAA1 in maintaining an acidic periplasm during SNF is supported by the observation that the expression of the respective gene is upregulated both upon the onset of nitrogen fixation and at alkaline pH values in freeliving bacteria. Similarly, the expression of the H. pylori ␣-CA was shown to be pH regulated by the ArsRS two-component system (43). In the CAA1::pFUS2 nodules, the formation of NH 4 ϩ outside the bacteroid inner membrane could result in a localized pH increase, which in turn would reduce the rate of NH 3 protonation, diffusion from the bacteroid cytosol, and transport to the plant cytosol, resulting in the observed increase in nodule N content.…”

“…Finally, a periplasmic ␣-type CA from H. pylori has been shown to be upregulated by low environmental pH, being essential for buffering the periplasmic acidic pH. Recent studies suggest that this CA is an integral component of the acid acclimation response that allows these neutralophile bacteria to colonize the stomach (24,43).…”

Posttranscriptional Regulation of Flagellin Synthesis in Helicobacter pylori by the RpoN Chaperone HP0958

“…Previous attempts to complement an HP0958 mutant by expressing the gene under the control of the ureA promoter were unsuccessful, suggesting that the overexpression of HP0958 interferes with its normal function (42). To eliminate the possible involvement of second-site mutations in our HP0958 mutant, we attempted to complement the mutation by expressing the HP0958 gene under the control of three well-characterized promoters, controlling the expression of flhA (a component of the flagellar apparatus), HP1186 (alpha-carbonic anhydrase) (60), and ahpC (alkyl hydroperoxide reductase) (34). The complementation plasmids were each introduced into a derivative of the P79 strain in which the HP0958 gene had been insertionally inactivated.…”

Novel Insights into Fur Regulation in Helicobacter pylori

“…However, despite the lack of regulatory factors, H. pylori must still be able to rapidly and efficiently respond to changes in its environment. As a result, the regulatory proteins utilized by this organism appear to function in extended capacities beyond what is typically seen for analogous proteins in other organisms (34,45,46,160,161,170). These differences in functionality can be seen in the expansion of the respective regulons to include genes that are not traditionally regulated by these factors in other bacteria, the ability of individual regulators to respond to multiple environmental conditions, and the ability to regulate genes using mechanisms not commonly seen in other bacterial species (34,45,46,160,161,170 …”