Identification of genes associated with natural competence in Helicobacter pylori by transposon shuttle random mutagenesis.
To identify genes involved in DNA transformation, we generated 1500 insertion mutants of a Helicobacter pylori strain by transposon shuttle mutagenesis. All mutant strains were screened for their frequency of natural transformation. A total of 20 mutant strains were found to exhibit a significantly decreased transformation frequency. DNA sequencing revealed seven genetic loci, including the reported comB locus, HP0017 (a putative virB4 homologue) and five loci without database match (HP0015, HP1089, HP1326, HP1424, and HP1473) from the 20 mutants. Reknockout of HP1326 revealed no impairment in natural transformation, while the other 5 mutants showed the same defective in natural transformation. Mutation of HP0017 severely impaired natural transformation both chromosome and plasmid DNA. Slot blot analysis revealed that some noncompetent strains had decreased virB4 RNA expression levels compared with competent strains. Nineteen ORFs had decreased expression levels in virB4 knockout mutant by microarray. Therefore, our data indicate that HP0017 is a virB4 homologue and is essential in the natural competence of H. pylori. HP0015, HP1089, HP1424, and HP1473 genes could be also involved in natural transformation.
Chang
K C
KC
Graduate Institute of Microbiology, National Taiwan University, College of Medicine, Taipei, Taiwan.
Yeh
Y C
YC
Lin
T L
TL
Wang
J T
JT
D015139:Blotting, Southern; D002876:Chromosomes, Bacterial / Q000235:genetics; D004251:DNA Transposable Elements / Q000235:genetics*; D030541:Databases, Genetic; D015536:Down-Regulation; D017353:Gene Deletion; D005798:Genes, Bacterial / Q000235:genetics*; D015698:Genomic Library; D016480:Helicobacter pylori / Q000235:genetics*; D016254:Mutagenesis, Insertional / Q000235:genetics / Q000379:methods*; D016366:Open Reading Frames / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D015183:Restriction Mapping; D014169:Transformation, Bacterial / Q000235:genetics*
D004251:DNA Transposable Elements