It is generally believed that most tissues acquire vitamin A as retinol delivered by its specific serum transport protein, retinol-binding protein (RBP). RBP is the sole transport protein for retinol in the circulation. Retinol delivered by RBP is oxidized within cells to retinoic acid, which regulates vitamin A-responsive gene expression, or to retinaldehyde which is needed in vision. Thus, RBP is assumed to play an essential role in mediating the biological actions of vitamin A. However, there are several RBP-independent pathways through which cells might acquire needed retinoid (vitamin A and its analogs). These include the direct delivery of retinoic acid through the circulation, the delivery of retinyl esters in chylomicrons and possibly other lipoproteins, the uptake of fully water-soluble glucuronides of both retinol and retinoic acid from the circulation, and through the uptake and conversion of carotenoids present in the circulation to retinoids. Although these delivery pathways are usually considered to be of minor impotence, our studies of the transthyretin (TTR)-null mouse suggest that this notion may not be correct. These mutant mice, which are fully healthy and fertile, have very low levels of plasma retinol-RBP, levels that are comparable to those of an extremely vitamin A-deficient animal. These studies imply that RBP may not be essential for maintaining the health of the organism. The goal of this proposal is to generate RBP- defective mice and to use this animal to explore the physiological role of serum RBP and to determine if RBP plays an indispensable role in any cellular process. We will generate an RBP-defective mouse by targeted disruption of the rbp gene, using the ES cell technology we previously employed to create the TTR-null mouse strain. We plan two strategies. The first will generate mice that totally lack RBP; the second will yield animals unable to synthesize RBP in the liver or adipose tissue (the two major sites of RBP synthesis). These mutants will be used for a detailed study of the uptake, transport and metabolism of retinoid under conditions of RBP deficiency. The specific aims of our study are: Aim 1. To generate RBP-deficient mice by targeted integration and to characterize these animals phenotypically and Aim 2. To investigate retinoid uptake from the diet and its delivery to tissues in these mutant mice.