Oncogene (2008) 27, 7047–7054 & 2008 Macmillan Publishers Limited All rights reserved 0950-9232/08 $32.00 www.nature.com/onc REVIEW Linking epithelial polarity and carcinogenesis by multitasking Helicobacter pylori virulence factor CagA M Hatakeyama Division of Molecular Oncology, Institute for Genetic Medicine, Hokkaido University, Sapporo, Japan Loss of cell polarity and tissue architecture is a hallmark second and fourth most common form of malignant of carcinomas that arise from epithelial cells. Recent tumors in male and female individuals, respectively, studies on Drosophila tumor suppressors have provided accounting for 876 000 estimated new cases and 405 000 evidence that epithelial polarity and cell proliferation are estimated deaths in the year 2000 (Parkin et al., 2001). functionally coupled, suggesting a function for polarity There isaccumulating evidence that chronic infection defects in the development of carcinomas. This notion is with Helicobacter pylori, a spiral-shaped bacterium that supported by the findings that mammalian orthologs of colonizes the gastric mucosa of more than half of the these Drosophila tumor suppressors are targeted by a world’shuman population, hasa causativefunction in number of viral oncoproteins. Chronic infection with the development of gastric carcinoma (Nomura et al., Helicobacter pylori is causally associated with gastric 1991; Parsonnet et al., 1991; Uemura et al., 2001). carcinoma. H. pylori virulence factor CagA (cytotoxin- Studieson H. pylori–gastric epithelial cell interactions associated gene A), which is delivered into gastric have greatly contributed to our current understanding epithelial cells through a bacterial type IV secretion of H. pylori-triggered mucosal lesions that direct gastric system, has an important function in cell transformation carcinogenesis, pointing to the H. pylori virulence factor through interacting with and deregulating SHP-2 phos- CagA (cytotoxin-associated gene A) as a central player phatase, a bona fide oncoprotein that is associated with in this process (Peek and Blaser, 2002; Hatakeyama, human malignancies. Recent studies have further revealed 2004, 2008). that CagA specifically binds and inhibits PAR1/MARK polarity-regulating kinase, thereby causing junctional and polarity defects in epithelial cells. Thus, the bacterial oncoprotein simultaneously targets the polarity-regulating Translocation of H. pylori CagA into gastric epithelial system and growth-regulatory system. These findings cells indicate that loss of cell polarity underlies the abnormal proliferation of epithelial cells that directs carcinogenesis. CagA isa 120 B145-kDa H. pylori protein that is Oncogene (2008) 27, 7047–7054; doi:10.1038/onc.2008.353 encoded by the cagA gene (Covacci et al., 1993; Tummuru et al., 1993). The cagA gene islocalized at Keywords: Helicobacter pylori; CagA; gastric carcino- one end of the cag pathogenicity island (cag PAI), a ma; epithelial polarity; PAR1/MARK 40-kb DNA segment that is considered to be horizon- tally transferred into the H. pylori genome (Censini et al., 1996; Akopyants et al., 1998). Whereasalmostall of the East Asian H. pylori isolates are cagA-positive, approximately half of the H. pylori strains isolated in Introduction Western countries do not carry cag PAI and thusare cagA negative. In addition to CagA, the cag PAI B Cell polarity isfundamental for cell-fate decisionduring contains 30 putative genes, among which 18 genes development aswell asfor the maintenance of differ- encode proteinsservingasbuilding blocksof a type IV entiated cells that constitute normal tissues. Tumors are secretion system. The type IV secretion system forms a formed by cells that have lost the ability to assemble and syringe-like structure that is capable of penetrating into create normal tissues. It is therefore reasonable to the cytoplasm of gastric epithelial cells. Infection with speculate that defects in cell polarity have an important cagA-positive H. pylori strains is associated with severe function in the development of tumors. The majority of mucosal inflammation that underlies peptic ulceration, human malignant tumorsthat arisefrom epithelial atrophic gastritis and gastric carcinoma (Blaser et al., tissues are termed carcinomas. Gastric carcinoma is the 1995; Kuipers et al., 1995; Parsonnet et al., 1997). Following the attachment of cagA-positive H. pylori to the surface of gastric epithelial cells, CagA is Correspondence: Dr M Hatakeyama, Division of Molecular Onco- delivered from the bacterium into the cytoplasm of host logy, Institute for Genetic Medicine, Hokkaido University, Kita-15, Nishi-7, Kita-ku, Sapporo 060-0815, Japan. cells through the type IV secretion system (Segal et al., E-mail: [email protected] 1999; Asahi et al., 2000; Backert et al., 2000; Odenbreit Linking epithelial polarity and carcinogenesis by H. pylori CagA M Hatakeyama 7048 et al., 2000; Stein et al., 2000). Thisprocessismediated Instead, they have a distinct EPIYA-containing at least partly through an interaction of H. pylori CagL sequence, termed the EPIYA-D segment, which is with integrins(Kwok et al., 2007). CagL, also encoded unique to East Asian CagA. by cag PAI, istargeted to the surfaceof the type IV secretion pili and acts as a specific adhesin that binds to the host integrin a5b1 in an Arg-Gly-Asp (RGD) motif- dependent manner. CagL–integrin interaction triggers Interaction of CagA with SHP-2 oncoprotein type IV injection of CagA into gastric epithelial cells. Within the host cells, CagA localizes to the inner surface Upon tyrosine phosphorylation, CagA acquires the of the plasma membrane, where it undergoes tyrosine ability to interact with the cytoplasmic protein tyrosine phosphorylation by Src family kinases or Abl kinase phosphatase SHP-2, which possesses two tandem- (Selbach et al., 2002; Stein et al., 2002; Poppe et al., repeated Src homology-2 (SH2) domainsat the 2007; Tammer et al., 2007). The tyrosine phosphoryla- N-terminal region (N-SH2 and C-SH2) and a protein tion site of CagA is characterized by the Glu-Pro-Ile- tyrosine phosphatase domain at the C-terminal region Tyr-Ala (EPIYA) motif, which ispresentin multiple (Higashi et al., 2002a, b) (Figure 1). Binding of tyrosine- numbersin the carboxy-terminal polymorphic region phosphorylated CagA to the SH2 domains causes a (EPIYA-repeat region) of the protein (Higashi et al., conformational change in SHP-2 that relievesintramo- 2002a, b). On the basis of sequences flanking the EPIYA lecular inhibition of the phosphatase domain by the N- motifs, four distinct EPIYA segments, EPIYA-A, -B, -C SH2 domain, resulting in aberrant activation of SHP-2 and -D, each of which containsa singleEPIYA phosphatase activity. Western and East Asian CagA motif, have been identified in the EPIYA-repeat species, respectively, bind SHP-2 through the tyrosine- region (Higashi et al., 2002b, 2005; Naito et al., 2006) phosphorylated EPIYA-C and EPIYA-D segments. (Figure 1). The representative CagA proteins of Western Notably, SHP-2-binding activity of the EPIYA-D H. pylori isolates (Western CagA) possess the EPIYA-A segment is significantly stronger than that of the and EPIYA-B segments followed by the EPIYA-C EPIYA-C segment, rendering East Asian CagA biolo- segment. Intriguingly, the EPIYA-C segment, consisting gically more active than Western CagA (Naito et al., of 34 amino-acid residues, variably multiplies in tandem, 2006). mostly from one time to three times, among different SHP-2 has an important function in signal transduc- Western CagA species as a result of homologous tion downstream of growth factor/cytokine receptors recombination of a 102-bp cagA gene segment encoding that promotescell proliferation, morphogenesisand EPIYA-C. The representative CagA proteins of H. motility (Neel et al., 2003). Consequently, expression of pylori isolated in East Asian countries (East Asian CagA in gastric epithelial cells causes morphological CagA) also possess the EPIYA-A and EPIYA-B transformation termed the hummingbird phenotype, segments but not the repeatable EPIYA-C segment. which ischaracterized by elongated cell shapewith SHP-2 N-SH2 C-SH2 Phosphatase domain ABC-CagA P P P A B C (Western) CM sequence PAR-1 Kinase domain ABD-CagA (East Asian) A B D CM sequence P P P SHP-2 N-SH2 C-SH2 Phosphatase domain Figure 1 Interaction of CagA with cellular targets. Western CagA contains EPIYA-A, EPIYA-B and EPIYA-C segments, whereas East Asian CagA comprises EPIYA-A, EPIYA-B and EPIYA-D segments. Each of the EPIYA segments contains a tyrosine-phosphorylatable EPIYA motif. Western and East Asian CagA interact with SHP-2 phosphatase in a tyrosine phosphorylation-dependent manner. The interaction involves the two SH2 domains of SHP-2 and tyrosine-phosphorylatable EPIYA-C or EPIYA-D segment of CagA. CagA also interacts with PAR1 kinase through the C-terminal 16-amino-acid residues termed the CM sequence, which locates immediately downstream of the EPIYA-C or EPIYA-D segment. CagA binds to the C-terminal 27-amino-acid residues present in the kinase catalytic domain of PAR1, thereby causing inhibition of the PAR kinase activity. CagA, cytotoxin-associated gene A; CM, CagA multimerization. Oncogene Linking epithelial polarity and carcinogenesis by H. pylori CagA M Hatakeyama 7049 dramatic cytoskeletal rearrangements (Segal et al., 1999; Disruption of tight junction and the loss of epithelial Higashi et al., 2002a). An important question is the polarity by CagA nature of the substrates that are dephosphorylated by CagA-activated SHP-2