Sequential Action ofTwo-Component Genetic Switches Regulates thePHO Regulon inBacillus subtilis

cells, anda mutation ineither genedecreased thetotal APaseexpressed underphosphate starvation conditions. Data presented hereshowthat aphoAphoBdouble mutantreduced APaseproduction during phosphate starvation by98%,indicating that these twogenesareresponsible formostoftheAPaseactivity during phosphate-limited growth. Thepromoter forphoA was cloned andused, withthephoBpromoter, toexamine phosphate regulation inB.subtilis. phoA-lacZ reporter gene assaysshowedthattheexpression ofthephoAgene commences as the culture enters stationary phase asaresult oflimiting phosphate concentrations inthegrowth medium, thereby mimicking thepattern oftotal APaseexpression. Induction persists forapproximately 2handisthenturned off.phoAistranscribed froma single promoterwhichinitiates transcription 19bpbefore thetranslation initiation codon. PhoPandPhoRaremembersofthetwo-component signal transduction systembelieved to regulate geneexpression inresponsetolimiting phosphate. Theexpression ofphoAorphoBinresponseto phosphate starvation was equally dependent on PhoPandPhoRforinduction. lacZ-promoter fusions showed thatbothphoAandphoBwerehyperinduced, orfailed toturnoffinduction after 2h,ina spoOAstrain ofB. subfilis. Mutations ingeneswhich arerequired forphosphorylation ofSpoOA, spoOBandspoOF, alsoresulted inphoAandphoBhyperinduction, suggesting thatphosphorylation ofSpoOAisrequired fortherepression of bothAPases inwild-type strains. Thehyperinduction ofeither APasegeneinaspoOAstrain was dependent on PhoPandPhoR.Analysis ofaphoP-lacZ promoter fusion showed thatthephoPRoperonishyperinduced ina spoOAmutantstrain, suggesting thatSpoOA-PrepressesAPasesbyrepressing phoPRtranscription. We proposea modelforPHO regulation inB.subtilis whereby thephoPRoperonistranscribed inresponseto limiting phosphate concentration, resulting inactivation ofthePHO regulon transcription, including transcription ofphoA andphoB. Whenthephosphate responsefails toovercomethenutrient deficiency, signals forphosphorylation ofSpoOAresult inproduction ofSpoOA-P, whichrepressestranscription ofphoPR, thereby repressing synthesis ofthePHO regulon. Nutrient depletion isa stress frequently encountered bya bacterium. A consequenceofnutrient depletion isthedisruption ofexponential growth asthecell enters stationary growth. During thistransition stage, geneswhichareinactive during exponential growth expressgeneproducts necessary forsurvival underthestress condition encountered. Depending on theenvironmental conditions, theBacillus subtilis cell may either entera nondividing stationary phase orbegin a developmental programleading totheformation ofa resistant endospore. Certainly, ifthegeneproducts expressed inresponsetothestress function toreplenish thedepleted nutrient before thecommitment tosporulation ismade,thecell can resumevegetative growth. Someofthefunctions expressed during thetransition stage