Abstract

Chronic infection with Helicobacter pylori cagA-positive strains is associated with atrophic gastritis, peptic ulceration, and gastric carcinoma. The cagA gene product, CagA, is delivered into gastric epithelial cells via type IV secretion, where it undergoes tyrosine phosphorylation at the EPIYA motifs. Tyrosine-phosphorylated CagA binds and aberrantly activates the oncogenic tyrosine phosphatase SHP2, which mediates induction of elongated cell morphology (hummingbird phenotype) that reflects CagA virulence. CagA also binds and inhibits the polarity-regulating kinase partitioning-defective 1 (PAR1)/microtubule affinity-regulating kinase (MARK) via the CagA multimerization (CM) sequence independently of tyrosine phosphorylation. Because PAR1 exists as a homodimer, two CagA proteins appear to be passively dimerized through complex formation with a PAR1 dimer in cells. Interestingly, a CagA mutant that lacks the CM sequence displays a reduced SHP2 binding activity and exhibits an attenuated ability to induce the hummingbird phenotype, indicating that the CagA-PAR1 interaction also influences the morphological transformation. Here we investigated the role of CagA dimerization in induction of the hummingbird phenotype with the use of a chemical dimerizer, coumermycin. We found that CagA dimerization markedly stabilizes the CagA-SHP2 complex and thereby potentiates SHP2 deregulation, causing an increase in the number of hummingbird cells. Protrusions of hummingbird cells induced by chemical dimerization of CagA are further elongated by simultaneous inhibition of PAR1. This study revealed a role of the CM sequence in amplifying the magnitude of SHP2 deregulation by CagA, which, in conjunction with the CM sequence-mediated inhibition of PAR1, evokes morphological transformation that reflects in vivo CagA virulence.

Highlights

  • H. pylori CagA oncoprotein is delivered into gastric epithelial cells, where it interacts with SHP2 oncoprotein and partitioning-defective 1 (PAR1) kinase

  • We demonstrated that CagA dimerization stabilizes CagA-SHP2 interaction and thereby potentiates deregulation of the SHP2 oncoprotein by CagA

  • Western CagA possesses at least two CagA multimerization (CM) sequences, each of which can independently bind to PAR1 [12]

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Summary

Background

H. pylori CagA oncoprotein is delivered into gastric epithelial cells, where it interacts with SHP2 oncoprotein and PAR1 kinase. Tyrosine-phosphorylated CagA binds and aberrantly activates the oncogenic tyrosine phosphatase SHP2, which mediates induction of elongated cell morphology (hummingbird phenotype) that reflects CagA virulence. CagA disrupts the tight junction and causes loss of epithelial cell polarity in a tyrosine phosphorylation-independent manner [11] This CagA activity is mediated by a specific interaction with partitioning-defective 1 (PAR1)/microtubule affinity-regulating kinase (MARK) [12]. CagA binds to all of the PAR1 family members in a tyrosine phosphorylation-independent manner and inhibits the kinase activity [14]. We demonstrated that CagA dimerization stabilizes CagA-SHP2 interaction and thereby potentiates deregulation of the SHP2 oncoprotein by CagA

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