prpR, ntrA, and ihf functions are required for expression of the prpBCDE operon, encoding enzymes that catabolize propionate in Salmonella enterica serovar typhimurium LT2.
The genes required for the catabolism of propionate in Salmonella enterica serovar Typhimurium are organized as two transcriptional units (prpR and prpBCDE) that are divergently transcribed from one another. Sequence homology to genes encoding members of the sigma-54 family of transcriptional activators and the identification of a consensus sigma-54 promoter 5' to the prpBCDE operon suggested that PrpR was required to activate expression of this operon. We isolated insertions in prpR and showed that prpR function was needed for growth on propionate as a carbon and energy source. A medium-copy-number plasmid carrying the lacZ gene under the control of the native sigma-54 promoter of prpBCDE was used to study prpBCDE operon expression. Transcription of the lacZ reporter in prpR, ntrA, and ihfB mutants was 85-, 83-, and 15-fold lower than the level of transcription measured in strains carrying the wild-type allele of the gene tested. These data indicated that PrpR, IHF, and transcription sigma factor RpoN were required for the expression of the prpBCDE operon. Further analysis of the involvement of the integration host factor (IHF) protein in the expression of this operon is required due to the well-documented pleiotropic effect the lack of this global regulator has on gene expression. Deletion of the 5' 615-bp portion of the prpR gene resulted in a PrpR(c) mutant protein that activated prpBCDE transcription regardless of the ability of the strain to synthesize 2-methylcitrate, the putative coactivator of PrpR. These results indicate that the N terminus of PrpR is the coactivator-sensing domain of the protein. When placed under the control of the arabinose-inducible promoter P(araBAD), expression of prpR(c) allele by arabinose had a strong negative effect on growth of the cell. It is proposed that this deleterious effect of PrpR(c) may be due to an uncontrolled ATPase activity of PrpR or to cross-activation of genes whose functions negatively affect cell growth under the conditions tested.
Palacios
S
S
Department of Bacteriology, University of Wisconsin-Madison, 53706-1567, USA.
Escalante-Semerena
J C
JC
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D003470:Culture Media; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D034982:Integration Host Factors; D008969:Molecular Sequence Data; D009154:Mutation; D009876:Operon; D010957:Plasmids / Q000235:genetics; D011422:Propionates / Q000378:metabolism*; D051877:RNA Polymerase Sigma 54; D012486:Salmonella typhimurium / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D017422:Sequence Analysis, DNA; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014161:Transduction, Genetic
D001426:Bacterial Proteins; D003470:Culture Media; D004268:DNA-Binding Proteins; D034982:Integration Host Factors; D011422:Propionates; D012808:Sigma Factor; D015534:Trans-Activators; C440508:integration host factor, Salmonella; D012321:DNA-Directed RNA Polymerases; D051877:RNA Polymerase Sigma 54; D000251:Adenosine Triphosphatases