Eukaryote polyphosphate kinases: is the 'Kornberg' complex ubiquitous?

Polyphosphate (poly P) is a polymer of up to several hundred phosphate residues and is important to a variety of cell processes. The main poly P synthetic enzyme in many bacteria is poly P kinase 1 (PPK1), which until recently had been detected among eukaryotes in some protists only. There is now evidence for the presence in several other eukaryotes of PPK1 homologues and also a second bacteria-type enzyme, PPK2. The latest genome databases reveal that the 'Kornberg' enzyme complex of three actin-related proteins, termed DdPPK2 in Dictyostelium discoideum, might also be ubiquitous in eukaryotes. Owing to the intimate association of poly P synthesis with the formation of structural fibres, this ubiquity indicates a central role for this molecule in the evolution of eukaryotic cells.

The long and short of it - polyphosphate, PPK and bacterial survival.

Inorganic polyphosphate (poly P) is present in all species tested to date, from each of the three kingdoms of life. Studied mainly in prokaryotes, poly P and its associated enzymes are important in diverse basic metabolism, in at least some structural functions and, notably, in stress responses. These numerous and unrelated roles for poly P are probably the consequence of its presence in life-forms from early in evolution. The genomes of many bacterial species, including pathogens, encode a homologue of a major poly P synthetic enzyme, poly P kinase 1 (PPK1). Loss of PPK1 results in reduced poly P levels, and deletion of the ppk1 gene in pathogens also results in a loss of virulence towards protozoa and animals. Thus far, no PPK1 homologue has been identified in higher-order eukaryotes and, therefore, PPK1 exhibits potential as a novel target for chemotherapy.

Interactions of some common pathogenic bacteria with Acanthamoeba polyphaga.

Protozoan grazing is a major trophic pathway whereby the biomass re-enters the food web. Nonetheless, not all bacteria are digested by protozoa and the number known to evade digestion, resulting in their environmental augmentation, is increasing. We investigated the interactions of Bacillus cereus, Enterococcus faecalis, Enteropathogenic Escherichia coli (EPEC), Listeria monocytogenes, Salmonella enterica serovar Typhimurium, and methicillin-sensitive Staphylococcus aureus (MSSA), with the amoeba, Acanthamoeba polyphaga. There was evidence of predation of all bacterial species except L. monocytogenes and S. aureus, where extracellular numbers were significantly higher when cultured with amoebae compared with growth in the absence of amoebae. Intracellular growth kinetic experiments and fluorescent confocal microscopy suggest that S. aureus survived and may even multiply within A. polyphaga, whereas there was no apparent intra-amoebal replication of L. monocytogenes and higher numbers were likely sustained on metabolic waste products released during coculture.

A polyphosphate kinase 1 (ppk1) mutant of Pseudomonas aeruginosa exhibits multiple ultrastructural and functional defects.

Pseudomonas aeruginosa, of medical, environmental, and industrial importance, depends on inorganic polyphosphate (poly P) for a wide range of functions, especially survival. Mutants of PAO1 lacking poly P kinase 1, PPK1, the enzyme responsible for most poly P synthesis in Escherichia coli and other bacteria, are defective in motility, quorum sensing, biofilm formation, and virulence. We describe here multiple defects in the ppk1 mutant PAOM5, including a striking compaction of the nucleoid, distortion of the cell envelope, lack of planktonic motility and exopolymer production, and susceptibility to the beta-lactam antibiotic carbenicillin as well as desiccation. We propose that P. aeruginosa with reduced poly P levels undergoes ultrastructural changes that contribute to profound deficiencies in cellular functions.

Amoebae promote persistence of epidemic strains of MRSA.

The control of healthcare-associated methicillin-resistant Staphylococcus aureus (MRSA) infection is of concern worldwide. Given the evidence that several pathogenic species replicate within amoebae and emerge more virulent and more resistant and the abundance of amoebae in healthcare settings, we investigated interactions of Acanthamoeba polyphaga with epidemic MRSA isolates. MRSA proliferated in the presence of amoebae, attributable partly to intracellular replication. Following 24 h of co-culture, confocal microscopy revealed that c. 50% amoebae had viable MRSA within phago-lysosomes and 2% of amoebae were heavily infected with viable cocci throughout the cytoplasm. Infection control strategies should recognize the contribution of protozoa.

Inorganic polyphosphate in Dictyostelium discoideum: influence on development, sporulation, and predation.

Dictyostelium discoideum, a social slime mold that forms fruiting bodies with spores, depends on inorganic polyphosphate (poly P) for its cycles of development and for nutritional predation on bacteria. The synthesis of poly P, a polymer of tens or hundreds of phosphate residues linked by high energy, ATP-like bonds, is catalyzed in most bacteria by poly P kinase (PPK1). The eukaryote D. discoideum possesses a homolog of PPK1. We report here that mutants of D. discoideum PPK1 (DdPPK1) have reduced levels of poly P and are deficient in development. Fruiting bodies are smaller and produce fewer spores, which appear to germinate like the wild type (WT). The DdPPK1 mutant formed smaller plaques on bacterial lawns compared with those of the WT. Predation by D. discoideum, assessed by uptake and digestion of Klebsiella aerogenes, showed that fewer bacteria were taken up by the DdPPK1 mutant compared with the WT and were killed less rapidly, indicating a role of poly P and/or DdPPK1 in phagocytosis. On Pseudomonas aeruginosa lawns, cleared plaques were observed with the bacterial PPK1 mutant but not with the WT P. aeruginosa. Thus, poly P is important in predation both for the predator and prey.

Inorganic polyphosphate in the origin and survival of species.

Inorganic polyphosphate (poly P), in chains of tens to hundreds of phosphate residues, linked by high-energy bonds, is environmentally ubiquitous and abundant. In prebiotic evolution it could have provided a flexible, polyanionic scaffold to assemble macromolecules. It has been conserved in every cell in nature. In prokaryotes, a major poly P synthetic enzyme is poly P kinase 1 (PPK1), which is found in 100 bacterial genomes, including numerous pathogens. Null mutants of PPK1, with low poly P levels, are defective in survival: namely, they show defective responses to physical/chemical stresses and predation. Pathogens with a PPK1 deletion are defective in biofilm formation, quorum sensing, general stress and stringent responses, motility, and other virulence properties. With the exception of Dictyostelium, PPK1 is absent in eukaryotes and provides a novel target for chemotherapy that would affect both virulence and susceptibility to antibacterial compounds. Remarkably, another PPK in Dictyostelium discoideum (PPK2) is an actin-related protein (Arp) complex that is polymerized into an actin-like filament, concurrent with its reversible synthesis of a poly P chain from ATP.