Interaction between the Helicobacter pylori CagA and alpha-Pix in gastric epithelial AGS cells.
The gastric pathogen Helicobacter pylori (H. pylori) translocates the CagA protein into epithelial cells by a type IV secretion process. Upon translocation into the host cell cytosol, CagA undergoes tyrosine phosphorylation. Phosphorylation of CagA occurs within the C terminus of the protein and is mediated by members of the Src family of tyrosine kinases. Phosphorylation of CagA induces the dephosphorylation of as yet unidentified cellular proteins, rearrangements of the host cell actin cytoskeleton, and cell scattering. This article aims to determine the cellular protein that interacts with CagA. Gastric epithelial AGS cells were stimulated with CagA-positive H. pylori (NCTC11637, at a bacteria/cell ratio of 500:1) and cultured in antibiotic-free medium. Proteins were isolated from the cells with or without H. pylori infection. CagA-interactive protein was determined by immunoprecipitation using anti-CagA antibody and proteomic analysis. We found that alpha-Pix interacts with CagA and alpha-Pix was constitutively expressed in AGS cells. Upon H. pylori stimulation, CagA was translocated into the cells and the expression of alpha-Pix (PAK-interactive exchange factor) was increased in AGS cells time dependently. The interaction of alpha-Pix with CagA was increased by H. pylori infection in AGS cells. Phosphorylation of CagA induces the dephosphorylation of alpha-Pix in AGS cells. alpha-Pix is a family of PAK-binding proteins that strongly activates PAK (p21-activated tyrosine kinase). PAK regulates changes in gene expression and mediates actin cytoskeletal and cell morphological changes. The novel finding of this study is that phosphorylation of CagA induces the dephosphorylation of alpha-Pix, which may modulate cytoskeletal changes of gastric epithelial cells through PAK.
Baek
Hye Yeon
HY
Department of Pharmacology and Institute of Gastroenterology, Yonsei University College of Medicine, Seoul, Korea.
Lim
Joo Weon
JW
Kim
Hyeyoung
H
D000942:Antigens, Bacterial / Q000378:metabolism / Q000502:physiology*; D001426:Bacterial Proteins / Q000378:metabolism / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000378:metabolism / Q000502:physiology*; D045744:Cell Line, Tumor; D003599:Cytoskeleton / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology*; D005753:Gastric Mucosa / Q000473:pathology; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism / Q000502:physiology*; D016481:Helicobacter Infections / Q000150:complications / Q000382:microbiology; D006801:Humans; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D040901:Proteomics; D064067:Rho Guanine Nucleotide Exchange Factors; D013274:Stomach Neoplasms / Q000378:metabolism / Q000382:microbiology*; D054462:p21-Activated Kinases
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D018797:Cell Cycle Proteins; D020662:Guanine Nucleotide Exchange Factors; D064067:Rho Guanine Nucleotide Exchange Factors; C080866:cagA protein, Helicobacter pylori; C515903:PAK1 protein, human; D017346:Protein Serine-Threonine Kinases; D054462:p21-Activated Kinases