Characterization of a modified lipoprotein of sporulation in Bacillus subtilis.

DIEZ VERÓNICA; SCHUJMAN GUSTAVO E.; DE MENDOZA DIEGO.

Rosario

Congreso; V Congreso Argentino de Microbiología General (SAMIGE); 2008

Sociedad Argentina de Microbiología General (SAMIGE)

In response to nutrient starvation and high cell density, a developmental program called sporulation is initiated in Bacillus subtilis. This process has become a paradigm of prokaryotic cell differentiation. An early event in this process is the formation of an asymmetric septum that divides the cell into two compartments, a smaller one, called prespore and a larger mother cell. The mother cell will assist in the prespore developmental process and will eventually lyse, releasing the dormant environmentally resistant spore. The differential behavior of each cell is generated by the activation of specific RNA-polymerase sigma factors in each compartment. The spores retain a specific sensory mechanism enabling them to germinate. The germination process is characterised by a series of degradative events leading to the loss of typical spore properties and outgrowth into a new vegetative cell. To identify hypothetical lipoproteins involved in sporulation, we labelled B. subtilis cultures with tritiated palmitic acid and found a putative lipoprotein expressed under the control of the mother cell specific sigma factor sE. We demonstrated that the protein detected is GerM, previously reported to be necessary for both spore formation and germination. A version of GerM mutated in its putative lipidation site (the Cys13 changed to Ala) called GerM1 was not able to incorporate tritiated palmitic acid. A mutant strain expressing gerM1 evidenced impaired sporulation efficiency. Measurements of sporulation parameters showed low levels of glucose dehydrogenase activity and dipicolinic acid content in sporulating gerM1 cells. A sporulation blockage was confirmed analyzing the expression profile of genes controlled by different sigma factors sequentially activated during the developmental process. Addition of germinants to the mutant spores resulted in the triggering of an incomplete germination with a normal loss of heat resistance (an early event) but significantly deficient in loss of spore refractility (a later event). These phenotypes are similar to those of a gerM null strain, suggesting that the lipobox site of this lipoprotein is critical for its proper function.