The Enterococcus faecalis SigV Protein Is an Extracytoplasmic Function Sigma Factor Contributing to Survival following Heat, Acid, and Ethanol Treatments

• DOI: 10.1128/jb.187.3.1022-1035.2005
• Publication Date: 2005-01-19T20:20:52Z
• Authors: Abdellah Benachour; Cécile Muller; Monika Dabrowski-...; Yoann Le Breton; Jean-Christophe G...; Alain Rincé; Yanick Auffray; Axel Hartke
• Keywords: 570; [SDV]Life Sciences [q-bio]; [SDV.BBM]Life Sciences [q-bio]/Biochemistry; Molecular Sequence Data; Restriction Mapping; Sigma Factor; Polymerase Chain Reaction; 03 medical and health sciences; Bacterial Proteins; STRESS RESPONSE; Enterococcus faecalis; [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology; Amino Acid Sequence; RNA, Messenger; Molecular Biology; DNA Primers; 2. Zero hunger; 0303 health sciences; SIGMA FACTOR; Base Sequence; Ethanol; Sequence Homology, Amino Acid; POLYMERASE; GENE EXPRESSION REGULATION; Genetic Complementation Test; Gene Expression Regulation, Bacterial; Hydrogen-Ion Concentration; 3. Good health; STRESS REGULON; Codon, Terminator; Mutagenesis, Site-Directed; Head; Sequence Alignment; Plasmids

ABSTRACT

Analysis of the genome sequence of Enterococcus faecalis allowed the identification of two genes whose protein products showed 33 and 34% identity with those of sigV and yrhM of Bacillus subtilis , respectively. These genes, named sigV and rsiV , are predicted to encode members of the extracytoplasmic function subfamily of eubacterial RNA polymerase sigma and anti-sigma factors, respectively. This group of sigma factors has been shown to regulate gene expression in response to stress conditions. sigV and rsiV were shown to be under the control of the same promoter. The transcriptional start site was determined, and the 1.5-kb mRNA transcript was shown to be overexpressed under glucose and complete starvation, as well as under physicochemical treatments. Three mutants, affected in sigV , rsiV , and both genes, were constructed by double-crossover recombination within the genome of E . faecalis strain JH2-2. Compared with the wild type and the rsiV mutant, the sigV mutants were more susceptible to heat shock, acid, and ethanol treatments and displayed decreased survival during long-term starvation. A nisin-inducible sigV gene construction used in complementation assays restored the wild phenotype of the sigV mutants, confirming the involvement of SigV in the heat shock, ethanol, and acid stress responses. Northern blot analysis carried out with the three mutant strains revealed the inhibition of sigV expression by the related anti-sigma factor gene rsiV . In addition, putative candidates of the sigV regulon determined by computer search for the sigV promoter sequence were analyzed.