| How do genetically identical progeny cells take on fates different from each other and from the parent cell from which they arose? Spore formation in the bacterium Bacillus subtilis is a tractable system with which to address this question. In response to nutrient depletion B. subtilis undergo a polar cell division that creates a smaller daughter cell, the forespore, and a larger daughter cell, the mother cell, that nurtures the forespore to maturity and then lyses. Immediately after polar division a transcription factor, sigmaF, that is present but inactive in the predivisional cell becomes active exclusively in the forespore. The activation of sigmaF sets up the forespore-specific pattern of gene expression and indirectly triggers the activation of a mother cell-specific transcription factor. Thus polar cell division and forespore-specific sigma F activation are the crucial events in establishing the developmental fates of both progeny cells. The goal of this thesis was to understand how the regulation of sigmaF activity is coupled to formation of the polar division septum.; sigmaF activity is regulated by SpoIIAB, an antisigma factor that binds to sigmaF and blocks its capacity to direct transcription; SpoIIAA, an anti-antisigma factor that activates sigma F by disrupting the AB-sigmaF complex; and SpoIIE, a septum-associated phosphatase that converts inactive phosphorylated SpoIIAA to its active, unphosphorylated form. sigmaF and all three of its regulatory proteins are present in the predivisional cell and in both daughter cells. However, sigmaF activity is restricted to the forespore after polar division. sigmaF activation is dependent on cell division, as mutants blocked in division are blocked in sigma F activation. My strategy for understanding the coupling of sigma F activation to cell division was to screen for mutants that activate sigma F independent of septum formation. Analysis of such mutants suggested that the concentration of unphosphorylated SpoIIAA must reach a threshold level to effect sigmaF activation. Further experiments provided evidence that SpoIIAA molecules below the threshold concentration are trapped in a complex with SpoIIAB. These findings lead to a model in which a threshold concentration of SpoIIAA cannot be reached until polar septation creates two compartments, each with different concentrations of the inhibitor and the activator of sigmaF.; This thesis also investigates the nature of the co-localization of SpoIIE with the cell division protein FtsZ, and SpoIIE's role in promoting polar versus medial division sites; the role of SpoIIE in insulating the sigma F regulatory pathway from cross talk from paralogous proteins; and the effect of a mutation in an ABC transporter on sigmaF activation. |
