Induced Helicobacter pylori vacuolating cytotoxin VacA expression after initial colonisation of human gastric epithelial cells.
To study the effect of initial colonisation on Helicobacter pylori gene expression, altered H. pylori gene transcription during co-culture with human gastric epithelial cells was determined. Therefore, an insertion library of H. pylori with random chromosomal fusions to a promoterless cat gene was grown in the presence of HM02 gastric epithelial cells and varying levels of chloramphenicol. One H. pylori transformant was chloramphenicol-resistant in the presence, but chloramphenicol-susceptible in the absence of gastric epithelial HM02 cells. This transformant had the promoterless cat gene inserted into the HP0887 gene, which encodes the vacuolating cytotoxin VacA, an important virulence factor of H. pylori. Reverse transcriptase polymerase chain reaction on cDNA of this transformant confirmed vacA upregulation near HM02 cells. These results show the applicability of this technique to study H. pylori gene regulation in its natural environment.
van Amsterdam
Karin
K
Academic Medical Center, Department of Medical Microbiology, P.O. Box 22660, 1100 DD Amsterdam, The Netherlands. k.vanamsterdam@amc.uva.nl
van Vliet
Arnoud H M
AH
Kusters
Johannes G
JG
Feller
Monique
M
Dankert
Jacob
J
van der Ende
Arie
A
D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D018920:Coculture Techniques; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004847:Epithelial Cells; D005753:Gastric Mucosa / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D016481:Helicobacter Infections / Q000378:metabolism* / Q000382:microbiology; D016480:Helicobacter pylori / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D012329:RNA, Bacterial / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013270:Stomach / Q000166:cytology / Q000382:microbiology*; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D012329:RNA, Bacterial; C074689:VacA protein, Helicobacter pylori; D037521:Virulence Factors; D015500:Chloramphenicol O-Acetyltransferase