Regulation of vitamin A synthesis from carotenoids

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DESCRIPTION (provided by applicant): All higher animals, including humans, must obtain vitamin A from the diet since they lack the ability to synthesize it de novo. Ultimately all vitamin A is formed from provitamin A carotenoids. The conversion process of provitamin A carotenoids to vitamin A has been studied for more than 4 decades although the cloning and purification of the enzyme responsible for the cleavage of provitamin A carotenoids to vitamin A were only recently accomplished when we and others cloned and characterized both mouse and human carotene cleavage enzyme (CCE). CCE was found to be ubiquitously expressed throughout the body and thus likely plays a general role in providing vitamin A to target tissues. The goal of our proposed studies is to understand the regulatory mechanisms controlling provitamin A conversion to vitamin A in intestine and liver. Two aspects of this regulation will be explored. First, we will examine the regulation of CCE activity by other proteins, possibly through direct protein-protein interactions. Cellular retinol-binding proteins (CRBPs) are known to interact with retinal, the product of carotene cleavage within cells. The effects of CRBPs on CCE activity will be examined using both in vitro and in vivo techniques. In addition, we will explore the direct down-stream enzyme to CCE, retinal reductase. We have identified two known short chain dehydrogenase/reductases, retSDR and RalR1, as candidate intestinal retinal reductases. The role of these two enzymes on retinol production from beta-carotene will be explored in depth. Finally, we will investigate the transcriptional regulation of hCCE expression. Here, we propose to examine the promoter of the human CCE gene to identify cis-regulatory elements and transcription factors that bind to these regulatory sequences in this gene. These data will provide insights into regulation of the CCE gene in humans. Taken together these studies will provide comprehensive understanding of CCE activity in two major tissue sites, intestine and liver.