Role of intracellular localization and phosphorylation of CagA in host cell activation.
Date Issued
2012
Date
2012
Author(s)
Hsu, Li-Ling
Abstract
Helicobacter pylori could cause gastritis, duodenal ulcer, and gastric cancer. CagA is regarded as a master virulent factor. CagA of H. pylori, a 120-145 kDa protein, is injected into gastric epithelial cells cytoplasm by type Ⅳ secretion system, which is accomplished by pilus and the cag PAI genes. Furthermore, the cytoplasmic CagA is phosphorylated by eukaryotic Src family kinase and Abl kinase. The phospho-CagA interacts with SHP-2 which becomes constitutive activated. The activated SHP-2 not only dephosphorylates focal adhesion kinase but also disrupts the E-cadherin and β-catenin pathway, leading to destroy the epithelial cell-cell junction. It is characterized by the spreading and elongated filopodia called hummingbird phenotype. The aberrant constitutive activation of SHP-2 is associated with tumor formation, therefore, CagA is considered as an oncoprotein. In the previous study, our lab firstly demonstrates that CagA could be directly translocated into human B lymphocytes from Helicobacter pylori. CagA is also phosphorylated in the cytoplasm, and then the phospho-CagA could bind to the phosphatase SHP-2. CagA activates p38, Erk, and anti-apoptosis molecules, Bcl-2 and Bcl-XL.
However, it is unclear about the intracellular location and the function of translocated CagA in B lymphocytes. In our previous study, it showed that CagA co-localized in nucleus in the immunohistochemistry staining of H. pylori+ mucosa-associated lymphoid tissue (MALT) lymphoma patients. Therefore, we want to study the intracellular location of CagA and to prove whether CagA would translocate into nucleus in B lymphocytes. We used B cells transfected with CagA plasmid or co-cultured with H. pylori, isolating the cytosolic and nuclear fraction. Our results indicated CagA could be detected in both cytoplasm and nucleus in B lymphocytes. Similarly, we also observed CagA could be translocated in nucleus of gastric epithelial cell lines, AGS cells. The expression of CagA is more abundant in nucleus than in the cytoplasm. Our results suggest that CagA may have an important role in the nucleus. By transfecting different truncated CagA plasmids into AGS cells, we reveal phosphorylation is not required for CagA nuclear translocation. CagA nuclear translocation is dependent on the fragment of CagA amine-terminus. We also isolated the nuclear fraction, and we demonstrated that nuclear translocated CagA co-immunoprecipitated with SHP-2. But the specific amine-terminal sequences for nuclear translocation and the role of CagA in nucleus is still needed to be investigated. Our results in this study will provide a crucial insight and contribution on CagA-induced B lymphocytes activation.
Subjects
H. pylori
phosphorylation
SDGs
Type
thesis
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