All-trans-retinoic acid (RA) is the bioactive derivative of vitamin A and ?-carotene that is essential for differentiation and development of embryonic tissues as well as the maintenance and robust performance of adult organs and tissues. In the nucleus, RA acts by binding to RXR/RAR heterodimeric transcription factors to regulate the expression of over 530 genes. RA is also known to have regulatory functions in the cytoplasm. During embryogenesis the levels of RA change in a strictly defined spatiotemporal pattern. Similarly, during adulthood the concentration of RA in various tissues and cells is maintained within narrow margins that are optimal for each type of cell. Disruption of RA homeostasis results in embryonic malformations, whereas in adult tissues aberrations in RA homeostasis can lead to pathophysiological changes that result in disease. Thus, it is critical to understand: (1) the molecular mechanisms whereby the cells maintain RA homeostasis; (2) how the cells adjust RA levels in response to varied physiological requirements; and (3) why these mechanisms fail in disease. Since the oxidation of retinol to retinaldehyde is the rate-limiting step in the pathway of RA biosynthesis that controls the overall rate of RA biosynthesis, it is important to identify and characterize the enzymes that catalyze this step and to understand the contribution of each enzyme to overall RA homeostasis. During the previous funding cycle, it was established that the baseline levels of RA in cells are maintained by a heterooligomeric retinoid oxidoreductase complex (ROC) formed by retinol dehydrogenase 10 (RDH10) and dehydrogenase/reductase 3 (DHRS3). Data from this and other laboratories indicate that RDH10 is also the primary enzyme responsible for the oxidation of retinol to retinaldehyde during early stages of embryogenesis. However, other yet unidentified retinol dehydrogenases appear to be more important in adult tissues. Preliminary data from this laboratory indicate that mice with a double knockout of genes encoding retinol dehydrogenase epidermal 2 (RDHE2) and RDHE2-similar (RDHE2S), display a phenotype consistent with reduced RA signaling in skin pilosebaceous unit and meibomian glands of eyelids. The data also suggest that the expression of RDHE2 and RDHE2S oscillates in a diurnal pattern and during various stages of hair follicle regeneration. We hypothesize that these inducible and RA-sensitive enzymes are responsible for the fluctuations of RA during the cycle of hair follicle regeneration, and for fine-tuning of the baseline RA levels, established by the ROC, in response to varied physiological requirements or pathophysiological conditions. To test this hypothesis, we will use the novel mouse models and custom-made antibodies generated during the previous cycle to determine the contribution of RDHE2/E2S to RA biosynthesis in skin and meibomian glands of the eyelid. The proposed studies will provide a comprehensive background to better understand the molecular mechanisms that maintain and/or disrupt RA homeostasis and will inform future strategies to develop targeted and more effective therapeutic interventions.
The correct cellular concentration of the bioactive vitamin A derivative, all-trans-retinoic acid (RA) is critical for optimal performance of adult organs and tissues. The enzymes called retinol dehydrogenases are important because they determine the rate of RA biosynthesis, but not all retinol dehydrogenases have yet been identified. The proposed studies will show whether the candidate enzymes, retinol dehydrogenase 2 and retinol dehydrogenase-similar, contribute to RA biosynthesis and homeostasis in mammals in vivo.
