O-antigen expression in Salmonella enterica serovar Typhi is regulated by nitrogen availability through RpoN-mediated transcriptional control of the rfaH gene.
The authors previously reported increased expression of the Salmonella enterica serovar Typhi (S. typhi) rfaH gene when the bacterial cells reach stationary phase. In this study, using a lacZ fusion to the rfaH promoter region, they demonstrate that growth-dependent regulation of rfaH expression occurs at the level of transcription initiation. It was also observed that production of the lipopolysaccharide (LPS) O-antigen by S. typhi Ty2 correlated with the differential expression of rfaH during bacterial growth. This was probably due to the increased cellular levels of RfaH, since expression of the distal gene in the O-antigen gene cluster of S. typhi Ty2, wbaP, was also increased during stationary growth, as demonstrated by RT-PCR analysis. Examination of the sequences upstream of the rfaH coding region revealed homologies to potential binding sites for the RcsB/RcsA dimer of the RcsC/YopJ/RcsB phosphorelay regulatory system and for the RpoN alternative sigma factor. The expression of the rfaH gene in rpoN and rcsB mutants of S. typhi Ty2 was measured. The results indicate that inactivation of rpoN, but not of rcsB, suppresses the growth-phase-dependent induction of rfaH expression. Furthermore, production of beta-galactosidase mediated by the rfaH-lacZ fusion increased approximately fourfold when bacteria were grown in a nitrogen-limited medium. Nitrogen limitation was also shown to increase the expression of the O-antigen by the wild-type S. typhi Ty2, as demonstrated by a similar electrophoretic profile to that observed during the stationary phase of growth in rich media. It is therefore concluded that the relationship between LPS production and nitrogen limitation parallels the pattern of rfaH regulation under the control of RpoN and is consistent with the idea that RpoN modulates LPS formation via its effect on rfaH gene expression during bacterial growth.
Bittner
Mauricio
M
Departamento de Bioquı́mica y Biologı́a Molecular, Facultad de Ciencias Quı́micas y Farmacéuticas, Universidad de Chile, PO Box 174, Correo 22, Santiago, Chile1.
Saldı As
Soledad
S
Departamento de Bioquı́mica y Biologı́a Molecular, Facultad de Ciencias Quı́micas y Farmacéuticas, Universidad de Chile, PO Box 174, Correo 22, Santiago, Chile1.
Estévez
Claudia
C
Departamento de Bioquı́mica y Biologı́a Molecular, Facultad de Ciencias Quı́micas y Farmacéuticas, Universidad de Chile, PO Box 174, Correo 22, Santiago, Chile1.
Zaldı Var
Mercedes
M
Departamento de Bioquı́mica y Biologı́a Molecular, Facultad de Ciencias Quı́micas y Farmacéuticas, Universidad de Chile, PO Box 174, Correo 22, Santiago, Chile1.
Marolda
Cristina L
CL
Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada N6A 5C12.
Valvano
Miguel A
MA
Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada N6A 5C12.
Contreras
Inés
I
Departamento de Bioquı́mica y Biologı́a Molecular, Facultad de Ciencias Quı́micas y Farmacéuticas, Universidad de Chile, PO Box 174, Correo 22, Santiago, Chile1.
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D003470:Culture Media; D004268:DNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D009584:Nitrogen / Q000378:metabolism*; D019081:O Antigens / Q000378:metabolism*; D010445:Peptide Elongation Factors / Q000235:genetics / Q000378:metabolism*; D051877:RNA Polymerase Sigma 54; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012485:Salmonella typhi / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D003470:Culture Media; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D019081:O Antigens; D010445:Peptide Elongation Factors; C106401:RfaH protein, E coli; D012808:Sigma Factor; D015534:Trans-Activators; C497751:rpoN protein, E coli; D012321:DNA-Directed RNA Polymerases; D051877:RNA Polymerase Sigma 54; D009584:Nitrogen