ABSTRACT
In bacteria, genes are often expressed from multiple promoters to allow for a greater spectrum of regulation. Transcription of rRNA genes in Escherichia coli uses two promoters, rrn P1 and rrn P2. Under the conditions examined previously, the P1 and P2 promoters were regulated in response to many of the same changes in nutritional conditions. We report here that rrn P2 promoters play unique roles in rRNA expression during transitional situations. rrn P2 promoters play a dominant role in rRNA synthesis as cells enter into and persist in stationary phase. rrn P2 promoters also play a role in the rapid increases in rRNA synthesis that occur during outgrowth from stationary phase and during the initial stages of rapid shifts to richer media. We demonstrate that rrnB P2 directly senses the concentrations of guanosine 5'-disphosphate 3'-diphosphate (ppGpp) and the initiating nucleoside triphosphate (iNTP), thereby accounting, at least in part, for the observed patterns of regulation. Our work significantly extends previous information about the regulators responsible for control of the rrn P2 promoters and the relationship between the tandem rRNA promoters.
Subject(s)
Escherichia coli/genetics , Gene Expression Regulation, Bacterial , Genes, rRNA , Promoter Regions, Genetic , RNA, Ribosomal , Base Sequence , Escherichia coli/metabolism , Nucleic Acid Conformation , Phosphates/chemistry , Phosphates/metabolism , RNA, Ribosomal/genetics , RNA, Ribosomal/metabolism , Transcription, GeneticABSTRACT
The mechanosensitive (MS) channels MscS and MscL are essential for the survival of hypoosmotic shock by Escherichia coli cells. We demonstrate that MscS and MscL are induced by osmotic stress and by entry into stationary phase. Reduced levels of MS proteins and reduced expression of mscL- and mscS-LacZ fusions in an rpoS mutant strain suggested that the RNA polymerase holoenzyme containing sigmaS is responsible, at least in part, for regulating production of MS channel proteins. Consistent with the model that the effect of sigmaS is direct, the MscS and MscL promoters both use RNA polymerase containing sigmaS in vitro. Conversely, clpP or rssB mutations, which cause enhanced levels of sigmaS, show increased MS channel protein synthesis. RpoS null mutants are sensitive to hypoosmotic shock upon entry into stationary phase. These data suggest that MscS and MscL are components of the RpoS regulon and play an important role in ensuring structural integrity in stationary phase bacteria.
Subject(s)
Bacterial Proteins/physiology , Escherichia coli Proteins/genetics , Escherichia coli/genetics , Gene Expression Regulation, Bacterial/genetics , Ion Channels/genetics , Ion Channels/physiology , Mechanotransduction, Cellular/physiology , Sigma Factor/physiology , Base Sequence , DNA Primers , Genes, Reporter , Mechanotransduction, Cellular/genetics , Plasmids/genetics , Transcription, Genetic , beta-Galactosidase/geneticsABSTRACT
The control of ribosomal RNA transcription is one of the most enduring issues in molecular microbiology, having been subjected to intense scrutiny for over 50 years. Rapid changes in rRNA expression occur during transitions in the bacterial growth cycle and following nutritional shifts during exponential growth. Genetic approaches and measurements of initiating nucleoside triphosphate (iNTP) and guanosine 5'-diphosphate, 3'-diphosphate (ppGpp) concentrations and of rRNA promoter activities showed that rapid changes in the concentrations of iNTPs and ppGpp account for the rapid changes in rRNA expression. The two regulatory signals are nonredundant: changes in iNTP concentration dominate regulation during outgrowth from stationary phase, whereas changes in ppGpp concentration are responsible for regulation following upshifts and downshifts during exponential phase. The results suggest a molecular logic for the use of two homeostatic regulatory mechanisms to monitor different aspects of ribosome activity and provide general insights into the nature of overlapping regulatory circuits.
Subject(s)
Bacteria/genetics , Gene Expression Regulation, Bacterial/genetics , RNA, Ribosomal/biosynthesis , RNA, Ribosomal/genetics , Transcription, Genetic/genetics , Bacteria/growth & development , Bacteria/metabolism , Dinucleoside Phosphates/genetics , Guanosine Tetraphosphate/genetics , Homeostasis/genetics , Promoter Regions, Genetic/genetics , Reaction Time/genetics , Ribosomes/genetics , Signal Transduction/geneticsABSTRACT
The seven rRNA operons in Escherichia coli each contain two promoters, rrn P1 and rrn P2. Most previous studies have focused on the rrn P1 promoters. Here we report a systematic analysis of the activity and regulation of the rrnB P2 promoter in order to define the intrinsic properties of rrn P2 promoters and to understand better their contributions to rRNA synthesis when they are in their natural setting downstream of rrn P1 promoters. In contrast to the conclusions reached in some previous studies, we find that rrnB P2 is regulated: it displays clear responses to amino acid availability (stringent control), rRNA gene dose (feedback control), and changes in growth rate (growth rate-dependent control). Stringent control of rrnB P2 requires the alarmone ppGpp, but growth rate-dependent control of rrnB P2 does not require ppGpp. The rrnB P2 core promoter sequence (-37 to +7) is sufficient to serve as the target for growth rate-dependent regulation.
