Figure 5. A, B. FipA is phosphorylated in vivo in a PknA-dependent manner.

A, B. M. smegmatis was transformed with a construct carrying pknA in antisense orientation under an acetamide-inducible promoter. Transformed cells were induced with 0.2% acetamide for the indicated periods of time in order to downregulate pknA expression. Antisensing was followed by immunoblotting of cell lysates with anti-PknA antibody, followed by reprobing with anti-FtsW antibody (A); or cell lysates at different periods of time were immunoprecipitated with anti-FipA antibody followed by immunoblotting with anti-phosphothreonine antibody and reprobing with anti-FipA antibody (B). Blots shown are representative of three separate experiments. IB: immunoblotting; IP: immunoprecipitation.