Preservation of Helicobacter pylori CagA Translocation and Host Cell Proinflammatory Responses in the Face of CagL Hypervariability at Amino Acid Residues 58/59

Abstract

Carriage of the CagA oncoprotein by the human gastric cancer-associated pathogen Helicobacter pylori is significantly associated with this typically benign chronic infection advancing to a potentially fatal outcome. However it remains to be elucidated why only a small subset of individuals infected with H. pylori CagA-positive strains develops gastric cancer. H. pylori translocates CagA into host cells using a type IV secretion apparatus that interacts with host integrin receptors via a three-amino-acid-residue RGD motif on the H. pylori protein CagL. The RGD motif of CagL also plays a major role in the induction of proinflammatory responses. Upstream of this motif is a conserved glycine flanked by four hypervariable amino acid residues (residues 58, 59, 61 and 62). Certain amino acid polymorphisms at 58 and 59 are significantly prevalent in strains from gastric cancer patients in particular geographic regions; Y58E59 is seen in Taiwan and D58K59 in India. In light of the seemingly contradictory findings of recent CagL mutagenesis studies, we have examined the contribution of sequence promiscuity specifically at CagL residues 58 and 59 to CagA translocation and H. pylori-mediated proinflammatory responses of gastric epithelial cells. Using isogenic mutants of H. pylori strains P12 and 26695 with amino acid substitutions at CagL residues 58 and 59, we determined that carriage of the polymorphisms Y58E59, D58K59, D58E59, N58E59 or N58K59 did not significantly alter the capacity of H. pylori to translocate CagA into, or induce IL-8 secretion in, host cells. Our findings, together with other recently published data, suggest that the variation at CagL residues 58 and 59 does not influence type IV secretion system function in isolation, but rather may work in concert with particular polymorphisms elsewhere in CagL to modulate disease progression.

Genetic diversity and amino acid sequence polymorphism in Helicobacter pylori CagL hypervariable motif and its association with virulence markers and gastroduodenal diseases

“…Based on previous studies, certain variants of CagLHM sequence containing five hypervariable amino acid residues (58, 59, 60, 61, and 62), which is located upstream of the RGD motif have been associated with gastric carcinogenesis . Recently, a global analysis of geographical diversity and polymorphism was carried out within the CagLHM motif of more than 500 amino acid sequences of CagL in gastric cancer‐associated H pylori isolates worldwide .…”

CagY-Dependent Regulation of Type IV Secretion in Helicobacter pylori Is Associated with Alterations in Integrin Binding

“…An RGD helper motif in CagL (FEANE) may also be important ( 13 ). However, other studies have failed to demonstrate CagL binding to β 1 integrins ( 12 ), have yielded discrepant results about the role of CagL polymorphisms ( 14 – 16 ), or have identified completely different integrin binding partners, including α V β 6 and α V β 8 ( 17 ). CagA, CagI, and CagY have also been shown to bind β 1 integrin using yeast two-hybrid, immunoprecipitation, and flow cytometry approaches ( 12 ).…”

Systematic site-directed mutagenesis of the Helicobacter pylori CagL protein of the Cag type IV secretion system identifies novel functional domains

“…This hypothesis cannot easily be verified in vitro . Transferring certain mutations in this region to a reference strain and testing these on host cells as has in part been done already46 might offer more clues as to whether these variations modulate the interaction of CagL with host cells. In addition, strain-specific structural differences in CagL variants, for example disulphide bridges, which can confer higher stability at lower pH42 are suggestive to be the result of functionally diversifying evolution.…”