The rfaH gene, which affects lipopolysaccharide synthesis in Salmonella enterica serovar Typhi, is differentially expressed during the bacterial growth phase

University of Santiago, Chile, CiudadSantiago, Santiago Metropolitan, Chile
FEMS Microbiology Letters (Impact Factor: 2.12). 11/2001; 204(1):123-8. DOI: 10.1111/j.1574-6968.2001.tb10874.x
Source: PubMed


We have cloned and sequenced the rfaH gene from Salmonella enterica serovar Typhi strain Ty2. The gene showed a high degree of similarity to the rfaH genes from Escherichia coli K-12 and S. enterica serovar Typhimurium. A rfaH mutant was constructed by site-directed mutagenesis. This mutant produced a rough lipopolysaccharide (LPS), with an incomplete core region. The defect in LPS expression that results from the rfaH mutation was corrected by a plasmid carrying the intact gene. The plasmid-borne rfaH gene also restored normal LPS synthesis in a rfaH mutant of E. coli. Reverse transcription-polymerase chain reaction analyses were performed to determine the effects of various environmental conditions on the expression of rfaH. The transcription of rfaH showed a growth-phase-dependent regulation, with maximal expression at the late exponential phase. Other environmental conditions, such as temperature or medium osmolarity, did not affect transcription of rfaH.

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    • "Interestingly, the level of rfaH mRNA was highly increased even in the early logarithmic growth phase (Fig. 3B). It was shown that transcription of rfaH in S. typhi is growth phase-dependent with peak expression at the end of the logarithmic phase (Rojas et al., 2001). This tight regulation seems to be altered by introducing multiple copies of rfaH through pRfaH. "
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    ABSTRACT: We reported earlier that the production of O antigen lipopolysaccharide (LPS) by Salmonella enterica serovar Typhi (Salmonella typhi) increases at the onset of stationary phase and correlates with a growth-regulated expression of the rfaH gene under the control of the alternative sigma factor RpoN (Microbiology 148 (2002) 3789). In this study, we demonstrate that RpoS also modulates rfaH promoter activity as revealed by the absence of growth-dependent regulation of an rfaH-lacZ transcriptional fusion and O antigen production in a S. typhi rpoS mutant. Introduction of a constitutively expressed rpoN gene into the rpoS mutant restored increased production of O antigen during stationary phase, suggesting that constitutive production of RpoN could overcome the RpoS defect. Similar results were observed when an rpoS rpoN double mutant was transformed with the intact rpoN gene. Thus, we conclude that both RpoS and RpoN control the rfaH promoter activity and concomitantly, the production of O-specific LPS in S. typhi.
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