Characterization of a novel inhibitory feedback of the anti‐anti‐sigma SpoIIAA on Spo0A activation during development in Bacillus subtilis

Abstract

SummaryCompartmentalized gene expression during sporulation is initiated after asymmetric division by cell-specific activation of the transcription factors s s s s F and s s s s E . Synthesis of these s s s s factors, and their regulatory proteins, requires the activation (phosphorylation) of Spo0A by the phosphorelay signalling system. We report here a novel regulatory function of the antianti-s s s s F SpoIIAA as inhibitor of Spo0A activation. This effect did not require s s s s F activity, and it was abolished by expression of the phosphorelay-independent form Spo0A-Sad67 indicating that SpoIIAA directly interfered with Spo0A ~ P generation. IPTG-directed synthesis of the SpoIIE phosphatase in a strain carrying a multicopy plasmid coding for SpoIIAA and its specific inhibitory kinase SpoIIAB blocked Spo0A activation suggesting that the active form of the inhibitor was SpoIIAA and not SpoIIAA-P. Furthermore, expression of the non-phosphorylatable mutant SpoIIAAS58A (SpoIIAA-like), but not SpoIIAAS58D (SpoIIAA-P-like), completely blocked Spo0A-dependent gene expression. Importantly, SpoIIAA expressed from the chromosome under the control of its normal spoIIA promoter showed the same negative effect regulated not only by SpoIIAB and SpoIIE but also by septum morphogenesis. These findings are discussed in relation to the potential contribution of this novel inhibitory feedback with the proper activation of s s s s F and s s s s E during development.

Compartmentalization of Gene Expression duringBacillus subtilisSpore Formation

“…Fujita and Losick propose that transient genetic asymmetry excludes important phosphorelay genes from the prespore at the time of asymmetric division. However, a recent study found that dephosphorylated SpoIIAA, thought to be found largely in the prespore (171), inhibits Spo0A-dependent transcription (6). This result suggests a supplemental or alternative mechanism to transient genetic asymmetry, that dephosphorylated SpoIIAA generates a feedback loop that prevents expression of Spo0A-dependent genes, such as spoIIG, from occurring in the prespore.…”

Synergistic effects of probiotic Leuconostoc mesenteroides and Bacillus subtilis in malted ragi (Eleucine corocana) food for antagonistic activity against V. cholerae and other beneficial properties

“…MTCC 5442 and Bacillus subtilis (Bs) natto RG4365 is obtained from Dr. Akira Nakamura, Tsukuba University, Japan (Arabolaza et al 2003;Lombardía et al 2006) The cultures were subcultured every week and incubated at 37°C overnight for growth. The cultures were stored at 4°C between transfers and were subcultured before experimental use.…”

TheBacillus subtilisSinR and RapA Developmental Regulators Are Responsible for Inhibition of Spore Development by Alcohol

“…Antibiotics were added to the media at the following concentrations for B. subtilis: 5 g/ml, chloramphenicol; 2 g/ml, kanamycin; 50 g/ml, spectinomycin; 20 g/ml, tetracycline; 0.5 g/ml, erythromycin; and 7 g/ml, neomycin. For determination of sporulation efficiency, ethanol-treated and nontreated cultures of B. subtilis were grown in SSM for 20 h and then serial dilutions were plated on solid SSM or LB before and after treatment with CHCl 3 (10% for 15 min) (1). ␤-Galactosidase activity in B. subtilis strains harboring lacZ fusions were assayed as described previously, and the specific activity is expressed in Miller units (1).…”

“…For determination of sporulation efficiency, ethanol-treated and nontreated cultures of B. subtilis were grown in SSM for 20 h and then serial dilutions were plated on solid SSM or LB before and after treatment with CHCl 3 (10% for 15 min) (1). ␤-Galactosidase activity in B. subtilis strains harboring lacZ fusions were assayed as described previously, and the specific activity is expressed in Miller units (1). The spore counting and ␤-galactosidase experiments for which results are shown in the figures were independently repeated five times, and a representative or average set of results is showed in each case.…”

“…At the onset of sporulation, cells divide asymmetrically into a large compartment (the mother cell) and a small compartment (the forespore) which are joined at the division septum. The polar division triggers cell-type-specific gene expression, first in the forespore and then in the mother cell (1,35,41).…”