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Molecular evolution of the VacA p55 binding domain of Helicobacter pylori in mestizos from a high gastric cancer region of Colombia.

ABSTRACT: The stomach bacterium Helicobacter pylori is one of the most prevalent pathogens in humans, closely linked with serious diseases such as gastric cancer. The microbe has been associated with its host for more than 100,000 years and escorted modern humans out of Africa. H. pylori is predominantly transmitted within families and dispersed globally, resulting in distinct phylogeographic patterns, which can be utilized to investigate migrations and bioturbation events in human history. Latin America was affected by several human migratory waves due to the Spanish colonisation that drastically changed the genetic load and composition of the bacteria and its host. Genetic evidence indicates that independent evolutionary lines of H. pylori have evolved in mestizos from Colombia and other countries in the region during more than 500 years since colonisation. The vacuolating cytotoxin VacA represents a major virulence factor of the pathogen comprising two domains, p33 and p55, the latter of which is essential for binding to the host epithelial cell. The evolution of the VacA toxin in Colombia has been strongly biased due to the effects of Spanish colonization. However, the variation patterns and microevolution of the p55 domain have not yet been described for this population. In the present study, we determined the genetic polymorphisms and deviations in the neutral model of molecular evolution in the p55 domain of 101 clinical H. pylori isolates collected in Bogotá, a city located in Andean mountains characterized by its high gastric cancer risk and its dominant mestizo population. The microevolutionary patterns of the p55 domain were shaped by recombination, purifying and episodic diversifying positive selection. Furthermore, amino acid positions 261 and 321 in the p55 domain of VacA show a high variability among mestizos clinical subsets, suggesting that natural selection in H. pylori may operate differentially in patients with different gastric diseases.

SUBMITTER: Gutierrez-Escobar AJ

PROVIDER: S-EPMC6507892 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Helicobacter pylori is a gram-negative bacterium that colonizes the human stomach and contributes to the development of gastric cancer and peptic ulcer disease. VacA, a toxin secreted by H. pylori, is comprised of two domains, designated p33 and p55. Analysis of the crystal structure of the p55 domain indicated that its structure is predominantly a right-handed parallel beta-helix, which is a characteristic of autotransporter passenger domains. Substitution mutations of specific amino acids within the p33 domain abrogate VacA activity, but thus far, it has been difficult to identify small inactivating mutations within the p55 domain. Therefore, we hypothesized that large portions of the p55 domain might be non-essential for vacuolating toxin activity. To test this hypothesis, we introduced eight deletion mutations (each corresponding to a single coil within a beta-helical segment spanning VacA amino acids 433-628) into the H. pylori chromosomal vacA gene.All eight of the mutant VacA proteins were expressed by the corresponding H. pylori mutant strains and underwent proteolytic processing to yield ~85 kDa passenger domains. Three mutant proteins (VacA Delta484-504, Delta511-536, and Delta517-544) were secreted and induced vacuolation of mammalian cells, which indicated that these beta-helical coils were dispensable for vacuolating toxin activity. One mutant protein (VacA Delta433-461) exhibited reduced vacuolating toxin activity compared to wild-type VacA. Other mutant proteins, including those containing deletions near the carboxy-terminal end of the beta-helical region (amino acids Val559-Asn628), exhibited marked defects in secretion and increased susceptibility to proteolytic cleavage by trypsin, which suggested that these proteins were misfolded.These results indicate that within the beta-helical segment of the VacA p55 domain, there are regions of plasticity that tolerate alterations without detrimental effects on protein secretion or activity, as well as a carboxy-terminal region in which similar alterations result in protein misfolding and impaired secretion. We propose that non-essential beta-helical coils and a carboxy-terminal beta-helical segment required for proper protein folding and secretion are features shared by numerous autotransporter passenger domains.

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Molecular evolution of the VacA p55 binding domain of Helicobacter pylori in mestizos from a high gastric cancer region of Colombia.

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