This project continues to be concerned with how cells coordinately regulate the expression of their genome during growth. Focus continues on processes mediated by guanozine 3', 5' bis-pyrophosphate (ppGpp). Ribosomal RNA transcription is exemplary of ppGpp inhibitory activity. We have found that ribosomal transcripts are modified (anti-termination) so as to ignore normal termination signals. This alteration depends on a sequence near the ribosomal promoter region that is genetically and physiologically homologous to an equivalent modification in phage lambda. Special terminator regions (called super-terminators) can stop an anti-terminating transcript. We have isolated such a region and constructed a large set of sequenced overlapping deletions that should enable functional dissection of the region. We have also isolated two mutants result in specific defects in super-termination, that terminate normally. New mutants of the major enzyme responsible for ppGpp degradation, encoded by the spoT gene, have been isolated and exploited to yield ten-fold variation in ppGpp basal levels during balanced growth; previously ppGpp variations could be studied only during starvations. We find an inverse correlation between growth rate and ppGpp levels. The spoT gene has been sequenced and extragenic suppressors of its function isolated that are important in analysis of ppGpp synthesis and decay. We have both genetic and physiological indications that histidine operon expression typifies a process positively regulated by ppGpp at the promoter level. Two histidine regulatory mutations (his W333 and hisU1820) have been mapped to the genes for gyrase subunits (gyrA and gyrB) suggesting super-helicity of DNA templates is important for his operon regulation as well as for ribosomal operon transcription.
