RNA polymerase-DNA interactions at complex gene regulatory regions

Singh, Shivani Shatrughana (2014). RNA polymerase-DNA interactions at complex gene regulatory regions. University of Birmingham. Ph.D.

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Abstract

RNA polymerase (RNAP) \(\sigma\) factor must recognise and bind to specific DNA elements, usually AT-rich, in order to initiate transcription. At AT-rich regulatory regions or with more than one \(\sigma\) factor binding site; RNAP has to distinguish between different targets to initiate transcription correctly. At two regulatory regions: i) cbpA regulatory DNA with overlapping binding sites for \(\sigma\)70 and 38 associated RNAP and ii) regulatory region for ehxCABD operon with AT content of 71 %, I examined how correct RNAP binding is ensured.
For cbpA regulatory region it was found that the shared promoter spacer region played a key role. I identified a location in spacer region that differently affected overlapping cbpA promoters. The base change at this position is sensed by \(\sigma\)70 side chain R451. Alterations in spacer sequence modulate conformation, making it easier, or more difficult, for R451-DNA interactions. Using tethered particle motion analysis, DNA compaction properties of cbpA gene product; CbpA was measured.
ehxCABD regulatory region contains many sequences resembling \(\sigma\) factor binding elements. RNAP is capable of binding to the correct promoter elements in this region only in the presence of a chromosome folding protein, H-NS which binds AT-rich DNA. H-NS “coats” ehxCABD regulatory region and enables specific RNAP binding. Finally, many intragenic promoters within ehxCABD operon were identified. We thus propose that H-NS plays a role in silencing this pervasive intragenic transcription.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

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