Morris, Joanna Ruth; (1999) Studies on the MUC1 promoter. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The gene for human MUC1 is selectively expressed in simple epithelia lining ducts and glands of many organs and is highly expressed in many carcinomas. Results herein show that the MUC1 1.4 kb 5' region and cDNA are sufficient for faithful tissue specific expression, high expression levels in lactating mammary gland and in mammary tumours in vivo. Transient transfection experiments show that promoter sequences -119/-62 are sufficient to direct epithelial specific transcription when cloned adjacent to the consitutively active HSV TK promoter. In vitro binding assays revealed that Sp1, Sp3, and a potentially novel factor, SpA, compete for DNA binding at the -97 GC box while USF and two less well characterised factors bind the sequence at -87/-75. The tissue-specific activity of the -119/-62 sequence is mediated by Sp1 and Sp3 binding activities, since a consensus Sp1 sequence cloned into the same TK-proximal site can substitute for the activity. Introduction of an Sp1 consensus elsewhere could not direct tissue specific activity, demonstrating that the position of the sequence is crucial. Transcription from the 1.4 kb MUC1 promoter reporter plasmid and the endogenous gene could be activated in non-epithelial cells by co-transfection of Sp1, suggesting that this factor is limiting in these cells. Functional analysis of a mutant promoter which lacked the ability to bind the SpA factor, showed an increase in transcription over wild-type. This was most marked in the epithelial cell line expressing least MUC1, MCF-7, suggesting that SpA may have a role in modulating MUC1 levels in epithelial cells. In support of this idea band-shift experiments showed that SpA levels were highest in MCF-7. These studies suggest that the binding of Sp1 and Sp3 and SpA to the -97 GC box may have an important role in both tissue-specific transcription and regulation of MUC1 expression in carcinomas.

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