Transcriptional and mutational analysis of the Helicobacter pylori urease promoter.

Urease is an essential virulence factor of the human gastric pathogen Helicobacter pylori, and is expressed to very high levels. The promoter of the urease operon contains sequences resembling the canonical -10 and extended -10 motifs, but no discernible -35 motif. To establish the role of different motifs and regions in the urease promoter, we fused the urease promoter to a genomic lacZ reporter gene in H. pylori, made substitutions in the aforementioned promoter motifs, and also made deletions in the upstream sequences removing regulatory sequences. Substitutions in the -10, extended -10 and predicted -35 motifs all significantly altered expression of the lacZ reporter gene, demonstrating their importance in transcription of the H. pylori urease operon. In contrast, sequential deletions upstream of the -35 region did not affect expression of the lacZ reporter gene. This demonstrates the modular structure of the H. pylori urease promoter, where basal levels of transcription are initiated from a typical sigma(70) promoter, which requires -10 and extended -10 motifs, and also its -35 motif for efficient transcription. Upstream sequences are not involved in basal levels of urease transcription, but play an important role in responses to environmental stimuli like nickel.

The stress-induced hsp12 gene shows genetic variation among Helicobacter pylori strains.

The transcriptional regulation, genetic variation and clinical relevance of the strain-specific hsp12 gene of the human gastric pathogen Helicobacter pylori were investigated. Although the transcription of the hsp12 gene in H. pylori strain 1061 was induced by growth under iron-, pH- and temperature-stress conditions, the gene was not essential for growth under these stress conditions. The locus containing the hsp12 gene showed considerable genetic variation. A total of eight different strain-specific alleles were identified, of which three are mosaic variants of the hsp12 gene and five that are unrelated to the hsp12 gene. The hsp12 locus of six paired sets of strains obtained from patients with 7-10-year time intervals remained unaltered, indicating that genetic variation does not occur during chronic infection. No significant association was found between the presence of a hsp12 gene and peptic ulcer disease in clinical isolates obtained from 26 patients. The stress-regulated, strain-specific hsp12 genes may be involved in adaptation of individual H. pylori strains to their specific hosts, and contribute to long-term colonization of the gastric niche.

Transcriptional phase variation of a type III restriction-modification system in Helicobacter pylori.

Phase variation is important in bacterial pathogenesis, since it generates antigenic variation for the evasion of immune responses and provides a strategy for quick adaptation to environmental changes. In this study, a Helicobacter pylori clone, designated MOD525, was identified that displayed phase-variable lacZ expression. The clone contained a transcriptional lacZ fusion in a putative type III DNA methyltransferase gene (mod, a homolog of the gene JHP1296 of strain J99), organized in an operon-like structure with a putative type III restriction endonuclease gene (res, a homolog of the gene JHP1297), located directly upstream of it. This putative type III restriction-modification system was common in H. pylori, as it was present in 15 out of 16 clinical isolates. Phase variation of the mod gene occurred at the transcriptional level both in clone MOD525 and in the parental H. pylori strain 1061. Further analysis showed that the res gene also displayed transcriptional phase variation and that it was cotranscribed with the mod gene. A homopolymeric cytosine tract (C tract) was present in the 5' coding region of the res gene. Length variation of this C tract caused the res open reading frame (ORF) to shift in and out of frame, switching the res gene on and off at the translational level. Surprisingly, the presence of an intact res ORF was positively correlated with active transcription of the downstream mod gene. Moreover, the C tract was required for the occurrence of transcriptional phase variation. Our finding that translation and transcription are linked during phase variation through slipped-strand mispairing is new for H. pylori.