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Title: Transcription initiation in Streptomyces coelicolor A3(2)
Author: Humphrey, Laurence J.
ISNI:       0000 0004 5921 998X
Awarding Body: University of Sussex
Current Institution: University of Sussex
Date of Award: 2016
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Recent studies into the stringent response and the discovery of a number of RNA polymerase binding proteins suggests that the model for bacterial transcription initiation in Actinobacteria may differ from that in Escherichia coli. In E. coli, the alarmone ppGpp, together with DksA, binds to RNA polymerase to elicit the stringent response. However, the ppGpp binding site on RNA polymerase is not conserved in S. coelicolor, although the organism possesses a DksA homologue. Deletion of DksA did not affect the growth and development of S. coelicolor, although its overexpression stimulated antibiotic production. Evidence is presented that suggests that this occurs through binding to the RNA polymerase secondary channel. The biological role of this protein remains unknown. CarD and RbpA are two RNA polymerase­‐binding proteins present in all Actinobacteria, including S. coelicolor and M. tuberculosis. Both proteins are critical for growth and have been identified as transcriptional activators from σHrdB­‐dependent promoters in vitro. Here it was demonstrated that CarD and RbpA activate transcription from rRNA promoters with a poorly conserved ­‐35 element. Surprisingly it was also found that both proteins can inhibit transcription from synthetic promoters with highly conserved ­‐35 elements. Chromatin immunoprecipitation followed by high throughput sequencing (ChIP­‐seq) experiments revealed that CarD and RbpA are found exclusively at promoter regions. RbpA is localised only at promoters recognised by σHrdB, whereas CarD also co‐localises with the alternative sigma factor σR during oxidative stress indicating that it lacks RNA polymerase holoenzyme specificity. The sigma specificity of RbpA was tested by the generation of sigma mutants that were defective in binding. In vivo, in vitro and ChIP­‐seq data presented in this study suggest that CarD and RbpA have an overlapping role in transcription initiation at σHrdB­‐dependent promoters in S. coelicolor.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QD0415 Biochemistry ; QR0075 Bacteria