The in vivo examination of transcriptional control mechanisms in mammalian cells

[摘要] In the investigations described in this thesis I have examined variousmechanisms involved in transcriptional control. The first chapter is a generalexamination of the mechanisms used in transcriptional control. The secondchapter addresses issues associated with the selection of transcriptional startsite. This work shows that core promoter usage can be altered in a tissue specificand environmentally-responsive manner. The third chapter of thiswork describes a significant improvement in the technique of ligation mediatedPCR-aided in vivo footprinting and genomic sequencing. Thisimprovement in the quality of in vivo footprint data allows the pattern ofprotein : DNA interactions to be obtained with greater signal to noise and for alarger group of DNA sequences. The fourth chapter of this thesis uses these invivo footprinting techniques to investigate the mechanisms controlling thetranscription of the mouse interleukin-2 gene upon T cell stimulation. T cellstimulation was shown to result in the coordinated occupancy of a number ofmajor groove binding proteins to the previously unoccupied IL-2 regulatoryregion in vivo. Finally, the appendix describes in vivo footprints at thedifferentiated-muscle-specific promoter of the delta-subunit of the nicotinicacetylcholine receptor. Multiple protein : DNA interactions were seen at thepromoter both before and after muscle cell differentiation, suggesting thattranscriptional regulation of this gene occurs at the level of proteinreplacement or an alteration of the ability of the assembled proteins to activatetranscription.

[发布机构] University:California Institute of Technology;Department:Biology

[关键词] Biology