Within all bacteria are intricate signal transduction pathways which allow monitoring and """"""""sensing"""""""" of environmental stress. The ability to rapidly adapt to harsh environments, like those encountered within the human body, is essential to cell survival. Among eubacteria, an ancient and ubiquitous stress response, the """"""""stringent response"""""""", is fundamental to implementing a regulatory network responsible for adjusting, redirecting, and balancing gene expression patterns to insure proper cell adaptation. The stringent response is initiated by production of a signal-transducer molecule, guanosine 3', 5' (bis) pyrophosphate, ppGpp, which binds directly to RNA polymerase reprogramming its promoter selectivity. In Escherichia coli, the importance of the stringent response has now been established through the construction of the first mutant strain in which production of the signal molecule ppGpp has been abolished. The absence of ppGpp leads to a pleiotropic phenotype in which adaptation and stress-dependent survival are severely restricted. Most striking among the phenotypes imposed by the ppGppo condition are the inability of otherwise prototrophic bacteria to grow in the absence of amino acids and the rapid onset of cell death following entrance into stationary-phase or saturated growth conditions. These growth defects have been used to select suppressor mutants capable of re-establishing cell survival by re-implementing the stringent response in the absence of ppGpp. Over 200 distinct suppressor mutants have now been isolated. All map to either rpoB, rpoC and rpoD, the genes encoding the b and b' subunits of core RNA polymerase and the initiation factor, s70 , respectively. Among these mutants should be those which mimic or by-pass ppGpp-dependent function. My previous studies with s70 mutants have shown, that on ppGpp regulated promoters, the rate of promoter escape by mutant RNA polymerases is increased. This discovery has implicated the initial transcribed sequence as a potential new promoter element which discriminates """"""""stringently controlled"""""""" promoters. I propose here a systematic in vivo and in vitro analysis of both the b and b' mutant suppressors and of the initial transcription sequence in order to elucidate the nature of ppGpp-modified RNA polymerase-promoter interactions. In addition, these investigation promise to uncover aspects of transcription which are fundamental and wide ranging in nature.
