Mycobacterium smegmatis PknL knockout mutants

To study the role of PknL in mycobacteria, we constructed strains of Mycobacterium smegmatis lacking functional copies of pknL (MSMEG_4243) and the adjacent gene MSMEG_4242, encoding the putative transcriptional regulator. Surprisingly, the double knock-out strains formed both smooth and rough colonies. Although both morphotypes differed from the parent mc2155 in phenotypes associated with the composition of the outer section of the cell envelope, such as sliding motility and the ability to form biofilms, the differences were more marked in the smooth colony strains. Further investigation revealed that all the smooth mutants contained mutations or insertions that inactivated the gene encoding nucleoid-binding protein Lsr2. The smooth mutants had increased expression of the IS1096 transposase, TnpA and also MSMEG_4727, which is similar to PKS5. The rough pknL/4242 deleted mutants formed poor biofilms and showed decreased, aberrant sliding motility and both phenotypes were corrected by complementation with the two deleted genes.