Inorganic polyphosphate and the induction of rpoS expression

Inorganic polyphosphate [poly(P)] levels in Escherichia coli were reduced to barely detectable concentrations by expression of the plasmid-borne gene for a potent yeast exopolyphosphatase [poly(P)ase]. As a consequence, resistance to H2O2 was greatly diminished, particularly in katG (catalase HPI) mutants, implying a major role for the other catalase, the stationary-phase KatE (HPII), which is rpoS dependent. Resistance was restored to wild-type levels by complementation with plasmids expressing ppk, the gene for PPK [the polyphosphate kinase that generates poly(P)]. Induction of expression of both katE and rpoS (the stationary-phase σ factor) was prevented in cells in which the poly(P)ase was overproduced. Inasmuch as this inhibition by poly(P)ase did not affect the levels of the stringent-response guanosine nucleotides (pppGpp and ppGpp) and in view of the capacity of additional rpoS expression to suppress the poly(P)ase inhibition of katE expression, a role is proposed for poly(P) in inducing the expression of rpoS.

The endopolyphosphatase gene: Essential in Saccharomyces cerevisiae

Endopolyphosphatases (Ppn1) from yeast and animal cells hydrolyze inorganic polyphosphate (poly P) chains of many hundreds of phosphate residues into shorter lengths. The limit digest consists predominantly of chains of 60 (P60) and 3 (P3) Pi residues. Ppn1 of Saccharomyces cerevisiae, a homodimer of 35-kDa subunits (about 352-aa) is of vacuolar origin and requires the protease activation of a 75-kDa (674-aa) precursor polypeptide. The Ppn1 gene (PPN1) now has been cloned, sequenced, overexpressed, and deleted. That PPN1 encodes Ppn1 was verified by a 25-fold increase in Ppn1 when overexpressed under a GAL promoter and also by several peptide sequences that match exactly with sequences in a yeast genome ORF, the mutation of which abolishes Ppn1 activity. Null mutants in Ppn1 accumulate long-chain poly P and are defective in growth in minimal media. A double mutant of PPN1 and PPX1 (the gene encoding a potent exopolyphosphatase) loses viability rapidly in stationary phase. Whether this loss is a result of the excess of long-chain poly P or to the lack of shorter chains (i.e., poly P60 and P3) is unknown. Overexpression of the processed form of Ppn1 should provide a unique and powerful reagent to analyze poly P when the chain termini are unavailable to the actions of polyPase and poly P kinase.