Diverse Phenotypes Resulting from Polyphosphate Kinase Gene ( ppk1 ) Inactivation in Different Strains of Helicobacter pylori

Abstract

Connections among biochemical pathways should help buffer organisms against environmental stress and affect the pace and trajectory of genome evolution. To explore these ideas, we studied consequences of inactivating the gene for polyphosphate kinase 1 (ppk1) in strains of Helicobacter pylori, a genetically diverse gastric pathogen. The PPK1 enzyme catalyzes synthesis of inorganic polyphosphate (poly P), a reservoir of high-energy phosphate bonds with multiple roles. Prior analyses in less-fastidious microbes had implicated poly P in stress resistance, motility, and virulence. In our studies, ppk1 inactivation caused the expected near-complete absence of poly P (>250-fold decrease) but had phenotypic effects that differed markedly among unrelated strains: (i) poor initial growth on standard brain heart infusion agar (five of six strains tested); (ii) weakened colonization of mice (4 of 5 strains); (iii) reduced growth on Ham's F-12 agar, a nutritionally limiting medium (8 of 11 strains); (iv) heightened susceptibility to metronidazole (6 of 17 strains); and (v) decreased motility in soft agar (1 of 13 strains). Complementation tests confirmed that the lack of growth of one ⌬ppk1 strain on F-12 agar and the inability to colonize mice of another were each due to ppk1 inactivation. Thus, the importance of ppk1 to H. pylori differed among strains and the phenotypes monitored. We suggest that quantitative interactions, as seen here, are common among genes that affect metabolic pathways and that H. pylori's high genetic diversity makes it well suited for studies of such interactions, their underlying mechanisms, and their evolutionary consequences.Many biochemical pathways are connected, in that a given metabolite can be generated and/or consumed by any of several enzymes, and the flux along one pathway can be influenced by genetic, culture-related, or environmental factors that affect traffic along complementary or competing pathways (30, 38). The resulting complex networks of interaction constitute a major focus of the new discipline of systems biology (see, e.g., reference 24); may underlie many of the epistasis (gene-gene interaction), penetrance, and quantitative-trait phenomena that are of major importance in medical, agricultural, and evolutionary genetics (13, 31); and are likely to affect the specificity and vigor of infection and virulence of pathogens. Metabolic networks exhibit intriguing formal similarities to phenomena such as food webs in natural ecosystems, patterns of human interaction, and the routing of electricity in power grids (43).It is with this perspective that we have been studying how inactivation of the ppk1 gene, which encodes polyphosphate kinase, affects Helicobacter pylori

Helicobacter pylori Evolution: Lineage-Specific Adaptations in Homologs of Eukaryotic Sel1-Like Genes

“…Alternatively, because distributions of slr genes in the three sequenced H. pylori genomes vary, these expansions could have been more recent, especially given the ease with which gene duplications can arise [47,48], possibly facilitated in H. pylori by its lack of a MutHSL DNA repair system [49], and/or induced by reactive metabolites generated during infection [50]. In either case, evidence of functional divergence among H. pylori slr homologs driven by positive selection makes it appealing to imagine their products affecting traits important for H. pylori mucosal colonization.…”

Environmental phosphate differentially affects virulence phenotypes of uropathogenicEscherichia coliisolates causative of prostatitis

“…12,13 Inorganic polyphosphate (polyP) was shown to be critical for attributes such as motility, quorum sensing, biofilm formation, resistance to oxidative, osmotic, heat, nutritional and alkaline stress, and stationary-phase survival in several microorganisms. [14][15][16][17][18][19][20][21][22][23] PolyP is a linear chain of tens to many hundreds of phosphate residues linked by high-energy phosphoanhydride bonds. 14,24 PolyP is usually accumulated during exponential phase of growth, and degraded at the beginning of stationary phase.…”

Helicobacter pylori Adhesion to Carbohydrates

“…In our experience, all selected transformants contain the desired mutant allele in place of the original wild-type allele, unless the gene in question is essential for viability (Chalker et al, 2001;Tan and Berg, 2004;Tan et al, 2005). …”

“…Gene105 and gene104 are arbitrary designations for orfs in the H. pylori genome. For detailed design of primer sequences for use with erythromycin and other resistance markers, see Tan and Berg (2004) and Tan et al (2005). Agarose gel electrophoresis of products from construction of insertion/deletion allele diagrammed in Fig.…”