Date of Award
1994
Degree Type
Dissertation
Degree Name
Doctor of Philosophy
Abstract
The transcriptional control of the two genes rpoB and rpoC encoding the {dollar}\beta{dollar} and {dollar}\beta\sp\prime{dollar} subunits of RNA polymerase was examined in a systematic fashion in order to evaluate the contribution of such control to the autogenous regulation of RNA polymerase synthesis. Numerous transcriptional fusions were constructed by placing restriction fragments of the rplKaJLrpoBC gene cluster upstream of the lacZ reporter gene on {dollar}\lambda{dollar} phage vectors. Assaying {dollar}\beta{dollar}-galactosidase activity from monolysogens of the recombinant phage permitted a high resolution scan of the gene cluster for transcriptional controls. The results precisely quantitate the efficiencies of two strong promoters, rplKp and rplJp, a weak promoter rplLp, and a transcriptional attenuator rpoBa located between rplL and rpoB. From this data, the relative contribution of each transcriptional signal to the overall transcription of rpoBC was determined. It was found that almost all transcription initiated from the strong rplKp promoter continued into the downstream rplJLrpoB genes; that rplJp continues to initiate efficiently despite such transcription from the upstream promoter; and most surprisingly, that the efficiency of termination at the rpoBa attenuator depended on whether transcription was initiated from rplKp, rplJp, or both promoters. Further examination of the latter phenomenon revealed that the efficiency of rpoBa was related in an inverse fashion to the frequency of transcription initiated by promoters located upstream of it. When the frequency of transcription into the attenuator was decreased over a 40-fold range by replacing rplJp and rplKp with alternative promoters, readthrough at rpoBa increased from 19% to 61%. When regions located upstream of rpoBa are deleted or inverted, the modulation of rpoBa may be abrogated. In addition, sequence analysis of the region can identify a site with remarkable homology to the boxB motif of the {dollar}\lambda\ nutR{dollar} site. Such findings suggest the potential involvement of antitermination in the modulation of rpoBa function. Modulation of rpoBa, together with the evidence by others that the global frequency of transcription is influenced by the cellular concentration of RNA polymerase, suggests that rpoBa may play a role in the autogenous control of rpoBC expression.
Recommended Citation
Steward, Keith L., "Transcriptional Regulation And Autogenous Control Of The Genes For The Beta And Beta' Subunits Of Escherichia Coli Rna Polymerase" (1994). Digitized Theses. 2390.
https://ir.lib.uwo.ca/digitizedtheses/2390