Effect of the spoIIID mutation on mother cell lysis in Bacillus thuringiensis.

SpoIIID is a small, sequence-specific DNA-binding protein which can direct many genes' transcription and has an effect on spore formation in Bacillus subtilis. We investigated the role of SpoIIID in mother cell lysis in Bacillus thuringiensis. A β-galactosidase assay based on the promoter fusions with lacZ indicated that the sigK gene was positively regulated by SpoIIID and σ negatively regulated the expression of sigE. The spoIIID mutant strain exhibited no mother cell lysis in Schaeffer's sporulation medium (SSM) but did in ½ Luria-Bertani (LB) medium. cwlC is an essential hydrolase gene for mother cell lysis. Moreover, the expression of a P-lacZ fusion in spoIIID mutant was proved to be higher in ½ LB medium than in SSM. HD (ΔspoIIID)(ΔcwlC) mutant was obtained by knocking out the cwlC gene in HD(ΔspoIIID) and displayed no mother cell lysis in both SSM and ½ LB mediums. The deletion of spoIIID decreased the crystal protein production in HD73. The expression of P and P promoter fusions with lacZ gene in the acrystalliferous HD(ΔspoIIID) mutant showed similar activity to that in the acrystalliferous HD73 strain before T and slightly higher than that in the acrystalliferous HD73 after T. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that Cry1Ac production in HD(ΔspoIIID) directed by the P and P promoters was at a similar level as that in HD73 wild strain. Altogether, these results suggested that the spoIIID mutant with P or P promoters could be an alternative delivery system for cry gene expression with no mature spore formation and medium-dependent mother cell lysis.