Natural variant of the Helicobacter pylori CagA oncoprotein that lost the ability to interact with PAR1.
Bottom Line: In the present study, we investigated the biological activity of v225d CagA, an Amerindian CagA of H. pylori isolated from a Venezuelan Piaroa Amerindian subject, because the variant CagA does not possess a canonical CM sequence.We found that v225d CagA interacts with SHP2 but not PAR1b.Furthermore, SHP2-binding activity of v225d CagA was much lower than that of CagA of H. pylori isolated from Western countries (Western CagA). v225d CagA also displayed a reduced ability to induce the hummingbird phenotype than that of Western CagA.
Affiliation: Division of Microbiology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.Show MeSH
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Mentions: H. pylori v225d CagA has an EPIYA-A segment, an EPIYA-B segment and an unusual EPIYA segment comprising an EPIYA-D-like sequence lacking the proximal CM sequence (Fig.1a, orange) on the right side of the EPIYA motif and an EPIYA-C-like sequence (Fig.1a, yellow) on the right side of the EPIYA motif (EPIYA-D/CΔCM). Furthermore, the distal CM sequence of v225d CagA is interrupted by the insertion of the partial EPIYA-D/CΔCM (pD/CΔCM) segment (Fig.1a, red dotted square), giving rise to the generation of a left-side CM fragment (L-CM) and a right-side CM fragment (R-CM) (Fig.1a).19 Because such an interrupted CM sequence is quite unique and its function is unknown, we first investigated the PAR1b-binding activity of v225d CagA. To do so, COS-7 cells were co-transfected with a HA-tagged CagA expression vector and an Omni-tagged PAR1b expression vector. H. pylori NCTC11637 strain-derived CagA (ABCCC CagA), which has three repeats of the EPIYA-C segment and four CM sequences, was used as a positive control. Total cell lysates (TCL) prepared were immunoprecipitated with an anti-Omni antibody followed by immunoblotting with an anti-HA antibody. Whereas ABCCC CagA was efficiently co-immunoprecipitated with PAR1b, v225d CagA was not (Fig.2a). This result indicates that v225d CagA does not bind to PAR1b in cells, even when both proteins are overexpressed.
Affiliation: Division of Microbiology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.