Retinoids are a class of compounds which are potent therapeutic agents for a number of dermatological diseases. In order to improve treatment of these diseases, it is necessary to elucidate the mechanism whereby retinoids, such as all-trans-retinoic acid (RA), exert their anti- proliferative effects. The long term goal of these studies is, therefore, to determine how RA inhibits keratinocyte growth. An important action of RA is keratinocytes is to inhibit ornithine decarboxylase (OD?), an enzyme whose products, the polyamines, are necessary for cell growth. It is therefore proposed to determine if inhibition of ODC gene expression is a direct consequence of retinoid action and how this repression is exerted. These experiments re designed to test the hypothesis that RA suppresses ODC gene transcription through a RA receptor (RAR)-mediated mechanism. This will be examined by (1) determining the effects of RA and/or cycloheximide on the relative transcription rates of the human ODC gene using run-on transcription; (2) determining which RAR is expressed in keratinocytes and discriminating between the actions of the cellular RA binding protein and the alpha-, beta-, and gamma-RARs through the use of chimeric RAR/estrogen receptors that will be transfected into keratinocytes; (3) identifying cis- acting DNA elements that interact with trans-acting factors to cause RA- mediated changes in ODC gene transcription, using in vitro footprinting, gel retardation assays, and transfectin of isolated elements ligated to the thymidine kinase-beta-globin gene; and (4) isolation of cDNAs encoding non- RAR trans-acting factors which are implicated in RA action. Elucidation of the mechanisms whereby RA suppresses ODC gene expression will yield important information on how retinoids exert their anti-proliferative effects.
