Expression of CEACAM1 or CEACAM5 in AZ-521 cells restores the type IV secretion deficiency for translocation of CagA byHelicobacter pylori

Abstract

Helicobacter pylori represents an important pathogen involved in diseases ranging from gastritis, peptic ulceration, to gastric malignancies. Prominent virulence factors comprise the vacuolating cytotoxin VacA and the cytotoxin-associated genes pathogenicity island (cagPAI)-encoded type IV secretion system (T4SS). The T4SS effector protein CagA can be translocated into AGS and other gastric epithelial cells followed by phosphorylation through c-Src and c-Abl tyrosin kinases to hijack signalling networks. The duodenal cell line AZ-521 has been recently introduced as novel model system to investigate CagA delivery and phosphorylation in a VacA-dependent fashion. In contrast, we discovered that AZ-521 cells display a T4SS incompetence phenotype for CagA injection, which represents the first reported gastrointestinal cell line with a remarkable T4SS defect. We proposed that this deficiency may be due to an imbalanced coexpression of T4SS receptor integrin-β 1 or carcinoembryonic antigenrelated cell adhesion molecules (CEACAMs), which were described recently as novel H. pylori receptors. We demonstrate that AZ-521 cells readily express integrin-β 1 , but overexpression of integrin-β 1 constructs did not restore the T4SS defect. We further show that AZ-521 cells lack the expression of CEACAMs. We demonstrate that genetic introduction of either CEACAM1 or CEACAM5, but not CEACAM6, in AZ-521 cells is sufficient to permit injection and phosphorylation of CagA by H. pylori to degrees observed in the AGS cell model. Expression of CEACAM1 or CEACAM5 in infected AZ-521 cells was also accompanied by tyrosine dephosphorylation of the cytoskeletal proteins vinculin and cortactin, a hallmark of H. pylori-infected AGS cells.Our results suggest the existence of an integrin-β 1and CEACAM1-or CEACAM5dependent T4SS delivery pathway for CagA, which is clearly independent of VacA.The presence of two essential host protein receptors during infection with H. pylori represents a unique feature in the bacterial T4SS world. Further detailed investigation of these T4SS functions will help to better understand infection strategies by bacterial pathogens.

Cysteine Residues in Helicobacter pylori Adhesin HopQ Are Required for CEACAM–HopQ Interaction and Subsequent CagA Translocation

“…Studying the structural elements of HopQ-CEACAM interaction has allowed us to demonstrate a key and specific role for disulfide bonds in H. pylori virulence, but, more importantly, offered a broader view of the larger network of proteins that supports what initially seems a straightforward protein-protein interaction. Although HopQ has been proposed to interact with the T4SS [35], the impaired CagA-translocation observed in our study is a result of the binding defect induced by the mutations inserted. The clear importance of Dsb-like proteins to H. pylori infection, whether it be HopQ binding, CagA translocation or others, prompts further work to precisely identify their contributions and further our understanding of their roles.…”

Type IV secretion of Helicobacter pylori CagA into oral epithelial cells is prevented by the absence of CEACAM receptor expression

“…To elucidate why CagA was not delivered into the oral cells, we investigated whether these cell types expressed CEACAM receptors. Previous Western blotting studies had revealed that AGS cells expressed three T4SSrelevant CEACAM members, sized approximately 120, 180 and 90 kDa, corresponding to CEACAM members 1, 5 and 6, respectively [22,23,26]. Here, we subjected uninfected HN, CAL-27 and BHY cells to flow cytometry and used monoclonal antibodies specific for detection of each of these three CEACAM members (Fig.…”

“…Most of the known gastric epithelial cell lines can express CEACAM receptors and permit CagA injection [22][23][24][25][26]. However, whether CEACAM receptors play a role in bacterial colonization of the oral cavity has not been studied yet.…”

Fusobacterium nucleatum supresses anti-tumor immunity by activating CEACAM1

“…Activation of CEACAM1 by CEACAM1 leads to the inhibition of immune cell activities, whereas the Opa activation of CEACAM1 impairs normal maturation of immature dendritic cells, suppresses lymphocyte responses to activating stimuli, and also impairs phagocytic engulfment of the bacteria. 13 CEACAM1 also binds HopQ which enables CagA translocation [14][15][16] and also protects tumors from immune cell attack. 17 CEACAM1 is expressed on the surface of a wide variety of tumors, and is considered to be a specific biomarker associated with tumor progression, metastasis and poor prognosis.…”