Impact of structural polymorphism for the Helicobacter pylori CagA oncoprotein on binding to polarity-regulating kinase PAR1b.

Hiroko Nishikawa,Masanori Hatakeyama,Takeru Hayashi,Fumio Arisaka,Toshiya Senda

doi:10.1038/srep30031

Abstract

Chronic infection with cagA-positive Helicobacter pylori is the strongest risk factor for atrophic gastritis, peptic ulcers, and gastric cancer. CagA, the product of the cagA gene, is a bacterial oncoprotein, which, upon delivery into gastric epithelial cells, binds to and inhibits the polarity-regulating kinase, partitioning-defective 1b (PAR1b) [also known as microtubule affinity-regulating kinase 2 (MARK2)], via its CagA multimerization (CM) motif. The inhibition of PAR1b elicits junctional and polarity defects, rendering cells susceptible to oncogenesis. Notably, the polymorphism in the CM motif has been identified among geographic variants of CagA, differing in either the copy number or the sequence composition. In this study, through quantitative analysis of the complex formation between CagA and PAR1b, we found that several CagA species have acquired elevated PAR1b-binding activity via duplication of the CM motifs, while others have lost their PAR1b-binding activity. We also found that strength of CagA-PAR1b interaction was proportional to the degrees of stress fiber formation and tight junctional disruption by CagA in gastric epithelial cells. These results indicate that the CM polymorphism is a determinant for the magnitude of CagA-mediated deregulation of the cytoskeletal system and thereby possibly affects disease outcome of cagA-positive H. pylori infection, including gastric cancer.

Highlights

Helicobacter pylori is a spiral-shaped Gram-negative bacterium that colonizes approximately 50% of the world’s population and is the causative agent of gastrointestinal diseases such as atrophic gastritis and peptic ulcers1,2
We examined for the first time the role of CagA multimerization (CM) polymorphism in the CagA-partitioning-defective 1b (PAR1b) interaction through quantitative analysis and evaluated the impact of the CM polymorphism on the CagA action towards the actin cytoskeletal system to better understand the gastrointestinal pathogenesis caused by cagA-positive H. pylori
To ascertain that the interaction is reproducible in a reciprocal pull-down, the GST-CagA-2CMW construct was used to pull-down PAR1b [39–364]

Summary

Introduction

Helicobacter pylori is a spiral-shaped Gram-negative bacterium that colonizes approximately 50% of the world’s population and is the causative agent of gastrointestinal diseases such as atrophic gastritis and peptic ulcers. Manifestations associated with chronic H. pylori infection vary considerably among distinct geographic regions and these differences have been attributed at least in part to polymorphisms of the H. pylori virulence factors such as CagA, VacA, IceA, BabA, DupA, and OipA6,7 Among those factors, much attention has been given to the structure-function relationship of CagA because of its strong association with gastric cancer. Almost all of the H. pylori strains isolated in East Asian countries produce a different class of CagA, termed the East Asian CagA, which, like Western CagA, contains the EPIYA-A and EPIYA-B segments but, instead of the EPIYA-C segments, carries a distinct segment termed EPIYA-D2,19. East Asian CagA, in contrast, does not contain a CM motif in its EPIYA-D segment but carries a single East Asian CM (CME) motif, which differs from the CMW by 5 amino acid residues, immediately downstream of the EPIYA-D segment. We examined for the first time the role of CM polymorphism in the CagA-PAR1b interaction through quantitative analysis and evaluated the impact of the CM polymorphism on the CagA action towards the actin cytoskeletal system to better understand the gastrointestinal pathogenesis caused by cagA-positive H. pylori

Methods

Results

Conclusion