Influence of Bacillus subtilis phoR on cell wall anionic polymers

Abstract

In Bacillus subtilis the Pho regulon is controlled by a sensor and regulator protein pair, PhoR and PhoP, that respond to phosphate concentrations. To facilitate studies of the Pho regulon, a strain with an altered PhoR protein was isolated by in vitro mutagenesis. The mutation in this strain (phoR12) leads to the production of a PhoR sensor kinase that, unlike the wild-type, is functionally active in phosphate-replete conditions. The lesion in phoRl2 was shown to be a single base change that results in an Arg to Ser substitution in a region of PhoR that is highly conserved in histidine sensor kinases. While a phoR-negative mutant was unable to induce the synthesis of cell wall teichuronic acid under phosphate-limited conditions, the phoR72 mutant showed a relative increase in teichuronic acid and a decrease in teichoic acid, even under phosphate-replete conditions. The latter suggests that some or all of the genes required for teichuronic acid synthesis are members of the Pho reg ulon.Keywords : phosphate stress, histidine sensor kinase, response regulator, alkaline phosphatase, signal transduction INTRODUCTIONPhosphorus is a major growth-limiting substrate in natural environments such as soil (Hulett, 1993). When the growth of Bacillus subtilis becomes limited by the availability of phosphorus, a programmed series of responses is induced that can ultimately lead to sporulation. Among the earliest of these responses are processes that reduce the cell's requirement for phosphorus, increase its affinity for the uptake of phosphate and lead to the production of extracellular hydrolytic enzymes that allow the cell to recover inorganic phosphate from organic sources such as teichoic acids (Grant, 1979;Eymann et al., 1996). If these conservation and scavenging mechanisms are effective, the developmental pathway leading to sporulation is reversed and the organism resumes vegetative growth. B. subtilis responds to phosphate stress by the coordinated induction of approximately 20 genes (Eymann et al., 1996). Prominent among these are genes of the Pho regulon (Seki et al., 1987;Hulett, 1993; Hulett et al., 1994a, b) , whose products include alkaline phosphatases (APase) , alkaline phosphodiesterase (APDase) , a highaffinity phosphate transporter and, as reported in this paper, proteins involved in teichuronic acid synthesis. a two-component, environment-sensing, signal-transduction system (Seki et al., 1987(Seki et al., ,1988. The components of this system, PhoP and PhoR, are equivalent, respectively, to the PhoB and PhoR proteins of Escherichia coli (Tommassen etal., 1982;Makino et al., 1985Makino et al., ,1986. In E. coli, PhoR is a membrane-spanning sensor protein with histidine protein kinase (HPKase) and phosphatase (PTase) activities Scholten & Tommassen, 1993). Activation at low phosphate concentrations leads to the production of PhoR -P which, in turn, acts as a substrate for the activation of the response regulator PhoB, required for the induction of the Pho regulon (Stock et al., 1989;Parkinson & Kofoid, 199...

Comparison of PhoP binding to the tuaA promoter with PhoP binding to other Pho-regulon promoters establishes a Bacillus subtilis Pho core binding site

“…(1997) have shown that teichuronic acid synthesis during phosphate starvation is dependent on PhoP and PhoR, since teichuronic acid does not accumulate in either a phoP or a phoR mutant. In these mutants, the teichoic acid cell wall content does not drop to the same low level as it does in the parent strain when phosphate is depleted, suggesting that teichoic acid is the anionic polymer in cell walls of phoP and phoR mutants even when phosphate is depleted from the growth environment (Muller et al, 1997). At least part of the explanation for this phenomenon is that PhoP -P represses the synthesis of teichoic acid (Liu et al, 1998a) for these apparent differences in Pho-regulon promoter expression may be that low levels of PhoP are phosphorylated by other kinases or other phosphodonors in the phoR mutant and that the concentration of PhoP -P in the cell required for activation of Phoregulon promoters differs.…”

“…Several lines of evidence indicate that the genes responsible for teichuronic acid synthesis also belong to the Pho regulon and therefore are regulated by PhoP and PhoR. First, teichuronic acid synthesis is induced during phosphate starvation (Ellwood & Tempest, 1969) and secondly, a phoP and a phoR mutant are unable to synthesize teichuronic acid during phosphate starvation (Muller et al, 1997). However, how PhoP and PhoR regulate synthesis of teichuronic acid during phosphate starvation, directly or indirectly, remains unknown.…”

“…Under phosphate-starvation conditions, B. subtilis synthesizes teichoic acid rather than teichuronic acid (Ellwood & Tempest, 1969;Grant, 1979;Lang et al, 1982;Muller, et al, 1997). Teichoic acid, which contains ribitol or glycerol phosphate polymers, accounts for 15 % of cellular phosphate in cells grown in phosphatereplete media (Archibald et al, 1993).…”

Bacillus subtilis NhaC, an Na + /H + Antiporter, Influences Expression of the phoPR Operon and Production of Alkaline Phosphatases

“…When the growth of B. subtilis becomes limited by the availability of phosphate, genes of the Pho regulon are either activated or repressed (24). These genes include phoA, phoB, and phoD encoding alkaline phosphatases (APases) and a phosphodiesterase-APase (13,14); the pstSACB1B2 operon encoding a high-affinity phosphate transporter (34); the tuaABCDEFGH, tagAB, and tagDEF operons involved in teichuronic acid and teichoic acid synthesis (23,25,28); glpQ encoding a glycerophosphoryl diester phosphodiesterase involved in the hydrolysis of deacylated phospholipids (2); and two genes, ydhF and ykoL, of unknown function (2,35). The members of the Pho regulon are controlled by the interaction of at least three two-component signal transduction systems (13).…”

Regulatory interactions between the Pho and σB-dependent general stress regulons of Bacillus subtilis

“…Genes of the Pho regulon that are induced in response to phosphate starvation include the following : phoA and phoB, encoding alkaline phosphatases (APases) Hulett et al, 1990) ; phoD, encoding a phosphodiesterase\APase, which has a putative role in cell-wall teichoic acid turnover (Eder et al, 1996) ; the pstSACBABB operon, encoding a highaffinity phosphate transporter (Qi et al, 1997) ; glpQ, encoding a glycerophosphoryl diester phosphodiesterase involved in the hydrolysis of deacylated phospholipids (Antelmann et al, 2000) ; the phoPR and resABCDE operons, encoding two-component signal-transduction systems PhoP-PhoR and ResD-ResE Hulett, 1996 ;Nakano et al, 2000) ; and two genes, ydhF and ykoL, of unknown function (Antelmann et al, 2000 ;Robichon et al, 2000). In addition, the expression of the tagAB and tagDEF operons, involved in the synthesis of a phosphate-containing cell-wall polymer, polyglycerolteichoic acid, is repressed (Mu$ ller et al, 1997 ;, while the expression of the tuaABCDEFGH operon, responsible for the synthesis of teichuronic acid, a non-phosphate-containing polymer that replaces teichoic acid in the wall, is induced during phosphate starvation (Mu$ ller et al, 1997 ;Lahooti & Harwood, 1999). Genes of the Pho regulon are controlled by the interaction of at least three two-component signaltransduction systems (Hulett, 1996).…”