Molecular Genetics of Bacillus subtilis RNA Polymerase

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DESCRIPTION (provided by applicant): Exposure of bacteria to diverse growth-limiting stresses induces the synthesis of a common set of proteins that provides broad protection against future lethal challenges. This general stress response enhances survival in the natural environment, in fresh and processed foods, and in certain pathogenic interactions. Among Bacillus subtilis and related Gram positive pathogens, this response is governed by the sigmaB transcription factor. Loss of sigmaB function causes increased sensitivity to multiple stresses, including acid, heat, osmotic, and oxidative stress. Our long term objective is to understand this response using Bacillus subtilis as a model, beginning with the sensors that detect the different stresses, extending through the signal transduction network that conveys this information to sigmaB, and ending with the physiological role of the 200 or more genes under sigmaB control. Of these areas, most is known about the signal transduction network, which functions by a 'partner switching" mechanism in which formation of alternate protein complexes is controlled by serine and threonine phosphoiylation. This mechanism apears to be very ancient, very plastic, and widespread among the eubacteria. Here it activates sigmaB in response to two classes of stresses: (i) energy stress, including starvation for carbon, phosphate, or oxygen; and (ii) environmental stress, including acid, ethanol, heat, or salt stress. These two classes are conveyed to aB by independent upstream signaling pathways, each terminating with a differentially regulated PP2C phosphatase and converging on the two direct regulators of sigmaB, the RsbV anti-anti-a and the RsbW anti-a factor. The energy branch consists of the RsbP phosphatase (with a PAS domain important for signaling) and RsbQ, a protein of unknown function required for signaling. The environmental branch has at least nine regulators, all joining to activate the RsbU phosphatase. How energy or environmental signals enter their respective branches is unknown. Also poorly understood is how the genes in the sigmaB regulon contribute to stress resistance. Experiments using DNA arrays indicate that sigmaB controls only a few genes with a direct protective function and instead governs changes in metabolism and envelope function which may confer a passive resistance. Our three specific aims address the following questions: (1) How do energy stress signals activate their branch of the network; (2) How do environmental signals activate their branch; and (3) What are the physiological roles of newly identified members of the sigmaB regulon, particularly those that may be involved in downstream signaling?