Cholesterol depletion reduces Helicobacter pylori CagA translocation and CagA-induced responses in AGS cells.
Infection with Helicobacter pylori cagA-positive strains is associated with gastritis, ulcerations, and gastric cancer. CagA is translocated into infected epithelial cells by a type IV secretion system and can be tyrosine phosphorylated, inducing signal transduction and motogenic responses in epithelial cells. Cellular cholesterol, a vital component of the membrane, contributes to membrane dynamics and functions and is important in VacA intoxication and phagocyte evasion during H. pylori infection. In this investigation, we showed that cholesterol extraction by methyl-beta-cyclodextrin reduced the level of CagA translocation and phosphorylation. Confocal microscope visualization revealed that a significant portion of translocated CagA was colocalized with the raft marker GM1 and c-Src during infection. Moreover, GM1 was rapidly recruited into sites of bacterial attachment by live-cell imaging analysis. CagA and VacA were cofractionated with detergent-resistant membranes (DRMs), suggesting that the distribution of CagA and VacA is associated with rafts in infected cells. Upon cholesterol depletion, the distribution shifted to non-DRMs. Accordingly, the CagA-induced hummingbird phenotype and interleukin-8 induction were blocked by cholesterol depletion. Raft-disrupting agents did not influence bacterial adherence but did significantly reduce internalization activity in AGS cells. Together, these results suggest that delivery of CagA into epithelial cells by the bacterial type IV secretion system is mediated in a cholesterol-dependent manner.
Lai
Chih-Ho
CH
Institute of Molecular and Cellular Biology & Department of Life Sciences, National Tsing Hua University, Hsinchu, Taiwan 300.
Chang
Yun-Chieh
YC
Du
Shin-Yi
SY
Wang
Hung-Jung
HJ
Kuo
Chun-Hsien
CH
Fang
Shih-Hua
SH
Fu
Hua-Wen
HW
Lin
Hui-Hao
HH
Chiang
Ann-Shyn
AS
Wang
Wen-Ching
WC
D000942:Antigens, Bacterial / Q000378:metabolism*; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D002784:Cholesterol / Q000378:metabolism*; D015169:Colony Count, Microbial; D004847:Epithelial Cells / Q000276:immunology / Q000378:metabolism / Q000382:microbiology*; D005753:Gastric Mucosa / Q000166:cytology* / Q000382:microbiology; D016480:Helicobacter pylori / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity* / Q000502:physiology; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis; D021962:Membrane Microdomains / Q000378:metabolism*; D009154:Mutation
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D016209:Interleukin-8; C080866:cagA protein, Helicobacter pylori; D002784:Cholesterol