Searching for genes mediating autolysis and bacterial cell death in Bacillus subtilis

Autolysis in Bacillus subtilis can be induced by either natural bacterial cell development events or a variety of external factors. Autolysins are a group of murein hydrolases that are capable of digesting bacterial cell wall peptidoglyca and act as the ultimate executors of autolysis. Because of their suicidal nature and their important roles in cellular functions, to understand how bacteria control autolysin activity is of great importance for understanding bacteria cell death and survival.; To have an overview of cellular response to autolysis induction, transcriptional profile of oxacillin or starvation induced autolysis was investigated by whole-genome microarray. 28 genes were induced and 49 genes were repressed significantly in oxacillin induced autolysis, while 49 genes were induced and 45 genes were repressed significantly in starvation induced autolysis. Common transcriptome under either condition included 7 significantly induced genes and 1 gene repressed. Lack of common regulatory components in 2 transcriptional profiles indicates potential different mechanisms underlying these two autolysis processes.; Mutational analysis of major autolysin lytABC operon indicated lytA was essential for autolysis induced by both oxacillin and starvation, while lytB was dispensable for LytC function in oxacillin induced autolysis but required in starvation induced autolysis. Defective autolysis caused by deletion of individual genes could not be trans-complemented by inducible expression of lytA or lytB alone.; To search for additional genes mediating autolysis, a transposon insertion library was constructed in wild type or lytC deletion strain. Transposon insertions resulting in defective autolysis were mapped to gene fliP, fliQ or sigD within fla/che operon or lytB within lytABC operon in wild type strain. A transposon insertion in lytC mutant which caused uncharacteristic autolysis under room temperature as well as better survival at 4°C was mapped to gene pksM. Transposon insertions in gene fliP, fliQ, sigD or lytB affected autolysis by decreasing transcription of lytC. Role of pksM in autolysis remains unknown.; It still remains elusive how bacteria control the process of autolysis. However, the approach developed in this study has the potential to identify additional genes regulating autolysis in B. subtilis.