Eloranta, Jyrki J; (1995) Regulation of the human beta-interferon promoter. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Beta-interferon (IFN-[beta]) is secreted from virus-infected cells to protect neighbouring cells from subsequent infection. In uninduced cells, IFN-[beta] production is repressed, whereas upon induction expression reaches high levels. Expression of IFN-[beta] is controlled at the level of transcriptional initiation. The changes in transcription rates are governed by a promoter, the complexity of which reflects its stringent regulation. Previous studies have indicated that the preinduction repression of the IFN-[beta] promoter is maintained by sequence-specific transcriptional repressors, although their identity has remained elusive. In contrast, several proteins have been identified that produce increased transcriptional rates upon induction. In this work, we have studied several aspects in regulation of the IFN-[beta] promoter, with the emphasis on factors contributing to preinduction repression. We have identified candidate proteins that have properties of preinduction repressors: They bind to the genetically defined negative regulatory elements, and their DNA affinities are decreased upon induction. The transcription factor Oct-1 is one such candidate repressor. We have investigated the modifications that may lead to its affinity decrease. We have established that the Oct-1 protein levels remain unchanged upon induction. The DNA binding domain of Oct-1 can be phosphorylated by nuclear kinases, specifically by protein kinase A; however, this phosphorylation alone cannot account for the decrease in DNA affinity. We have established a transient transfection system, which allows the detection of preinduction repression of the promoter and, using this, studied the consequences of the modulation of intracellular levels of the Oct-1 protein. While we have obtained evidence that Oct-1 can have a repressive effect on the IFN-[beta] promoter, our analysis has also revealed an unexpected degree of complexity in the regulatory properties of Oct-1. The Un1 and Un2 complexes are further candidates for preinduction repressors. We have studied their DNA binding specificity and shown that their DNA binding can be regulated by their phosphorylation status. The results of the purification of the polypeptide components of Un1 and Un2 are presented. Finally, an analysis of the positive regulatory domain IV and an investigation into the involvement of protein kinase A in the signalling pathways leading to the induction of the promoter, are presented.

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