Expression of a Cryptic Secondary Sigma Factor Gene Unveils Natural Competence for DNA Transformation in Staphylococcus aureus
A) Positive selection tetracycline resistance (tet) reporter plasmid (pTet-rep). Reporter strains carrying pTet-rep were selected with 5 µg/ml tetracycline. cat: chloramphenicol resistance gene. bgaB: β-galactosidase gene. PcomG: promoter region of comG operon. PcomE: promoter region of comE operon. B) Fluctuation tests indicate that SigH activation occurs spontaneously. The y-axis represents the numbers of SigH active colonies detected from 109 cfu. Cells were grown in drug-free TSB and then selected for tetracycline resistance. Open symbols: aliquots from a single flask. Closed symbols: independent cultures in separate test tubes. Diamonds: N315 pTet-rep, Triangles: RNtet-rep (RN4220 pTet-rep). C) SigH activity can be stably maintained through generations without selection pressure. pTet-rep was cured from SigH active cells, and pRIT-PcomE-bgaB was introduced. Cells were grown on TSA plates containing 100 µg/ml X-gal and 12.5 µg/ml chloramphenicol. Rare white cells (about 1%) that lost SigH activity were also observed (see text).