SPECIFIC ORGANIZATION OF THE NEGATIVE RESPONSE ELEMENTS FOR RETINOIC ACID AND THYROID-HORMONE RECEPTORS IN KERATIN GENE FAMILY

Retinoic acid and thyroid hormone are important regulators of epiderma l growth, differentiation, and homeostasis, Retinoic acid is extensive ly used in the treatment of many epidermal disorders ranging from wrin kles to skin cancers. Retinoic acid and thyroid hormone directly contr ol the transcription of differentiation-specific genes including kerat ins. Their effect is mediated through nuclear receptors RAR and T3R. W e have previously identified the response element in the K14 gene, K14 RARE/TRE, to which these receptors bind, and found that it consists of a cluster of five half-sites with variable spacing and orientation. T o determine whether this specific structure is found in other keratin genes, we have mapped and analyzed the RARE/TRE elements in three addi tional epidermal keratin genes: K5, K6, and K17. We used three differe nt approaches to identify these elements: co-transfection of promoter deletion constructs, gel-shift assays, and site-specific mutagenesis. We localized the RARE/TRE elements relatively close to the TATA box in all three promoters. All three RARE/TRE elements have a similar struc tural organization: they consist of clusters of 3-6 half-sites with va riable spacing and orientation. This means that the clustered structur e of the RARE/TREs is a common characteristic for keratin genes. RARE and TRE in the K5 promoter are adjacent to each other whereas in the K 17 promoter they overlap. All three keratin REs bind specifically both RAR and T3R in gel-shift assays. Interestingly, addition of ligand to the receptor changes the binding pattern of the T3R from homodimer to monomer, reflecting the change in regulation from induction to inhibi tion.