Multistep activation of the Helicobacter pylori effector CagA

Abstract

Chronic infection with the Gram-negative bacterium Helicobacter pylori is a major risk factor for the development of gastric cancer. Accumulating evidence indicates that the H. pylori virulence determinant cytotoxin-associated gene A (CagA) has a key oncogenic role in the process. Certain biological activities of CagA require its tyrosine phosphorylation by host cell kinases. In this issue of the JCI, Mueller and colleagues report their detailed kinetic and functional analysis of CagA phosphorylation, which indicates that c-Src and c-Abl kinases sequentially phosphorylate CagA. Interestingly, the two phosphorylation events need not occur on the same CagA molecule but are both required for the biological effects of CagA. The results provide a clinically relevant example of how a successful bacterial pathogen has evolved to exploit the tightly coordinated, sequential activity of host cell kinases for virulence factor activation and induction of pathology.

Figure 1. Schematic representation of the most common Western and East Asian CagA variants.

A, B, C, and D segments are characterized by the upstream and downstream nucleotide sequences flanking the EPIYA motif. (A) Three commonly occurring Western CagA variants. (B) A common East Asian CagA variant. Additional EPIYA configurations found in Western CagA proteins include AB, AC, ACC, ACCC, ABCCCC, ABCCCCC, ABCABC, ABCBCC, ABABC, ABBBC, ABBCC, AABC, ABBC, BC, CC, and C. Other East Asian CagA proteins exhibit the configurations AABD, ABDD, ABBBD, ABDABD, ABABD, ABDBD, ABBD, BABD, AD, and BD.

Figure 2. Simplified model showing the consecutive phosphorylation of EPIYA-C, -A, and -B motifs by c-Src and c-Abl kinases in the commonly occurring Western CagA EPIYA-ABC configuration, and subsequent downstream effects.

Src family kinases become activated between 30 and 90 minutes after infection and exclusively phosphorylate the tyrosine residue of an EPIYA motif surrounded by C segment sequences. SHP-2 is known to bind to EPIYA-C^PY. EPIYA-C^PY CagA becomes a substrate for Abl family kinases at 90–180 minutes after infection; c-Abl is in principle capable of phosphorylating tyrosine residues in EPIYA motifs located in either A, B, or C segments, but never phosphorylates more than 2 residues per molecule (preferentially A and C). Triple-phosphorylated CagA is not detectable in infected cells. CagA^PY interactors such as CSK and PI3K bind to EPIYA-A^PY and EPIYA-B^PY motifs. Double-phosphorylated EPIYA-AC^PY CagA triggers the activation of several downstream signaling pathways; the Ras-Raf-MEK pathway is shown representatively. The biological and cell morphological consequences of CagA phosphorylation are cell elongation and increased cell motility. Single-phosphorylated EPIYA-C^PY CagA or double-phosphorylated EPIYA-BC^PY CagA do not efficiently induce Ras-Raf-MEK signaling. In East Asian CagA variants, EPIYA-D functionally replaces EPIYA-C as the initial motif to be phosphorylated by c-Src (not shown).