Optimization of a two-plasmid system for the identification of promoters recognized by RNA polymerase containing Staphylococcus aureus alternative sigma factor sigmaB.
We optimized a previously established two-plasmid system for the identification of Staphylococcus aureus promoters that are recognized by the alternative transcription factor sigma(B). The method allowed the identification of 18 S. aureus sigma(B)-dependent promoters, 12 of which are reported here for the first time to be sigma(B)-dependent. S1-nuclease mapping of the respective transcriptional start points revealed that all the promoters contained sequences exhibiting high similarity to the consensus sequence of Bacillus subtilis sigma(B)-dependent promoters. The promoters governed expression of genes encoding proteins proposed to be involved in various cellular functions, including the stress response genes and virulence-associated clfA gene for fibrinogen-binding clumping factor. Comparison of the nucleotide sequences upstream of the identified transcription start points identified a sigma(B) consensus promoter (GttTaa-N(12-15)-gGGTAt) that is highly homologous to that of sigma(B) of B. subtilis.
Homerova
Dagmar
D
Institute of Molecular Biology, Center of Excellence for Molecular Medicine, Slovak Academy of Sciences, Dubravska cesta 21, 845 51 Bratislava, Slovak Republic.
Bischoff
Markus
M
Dumolin
Alexis
A
Kormanec
Jan
J
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics; D008967:Molecular Biology / Q000379:methods*; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012808:Sigma Factor / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000201:enzymology / Q000235:genetics*
D001426:Bacterial Proteins; C079011:SigB protein, Bacteria; D012808:Sigma Factor; D012321:DNA-Directed RNA Polymerases