The essential role of vitamin B12 (Cbl) in recycling folate and thereby in DMA synthesis and single carbon metabolism and in the generation of methionine, the precursor for biological methylations is indicated by the clinical presentation of megaloblastic anemia and neurologic abnormalities in Cbl deficiency. The two metabolites, homocysteine (HCY) and methylmalonic acid (MMA) that is elevated in both Cbl and folate deficiency have been implicated in vascular damage and neurologic abnormalities respectively. The incidence of these elevated metabolites in the elderly population is a cause for concern. In the absence of dietary deficiency and malabsorption, the cellular uptake of Cbl is the likely cause of elevated HCY and MMA. Transcobalamin (TC), the Cbl transporter secreted by the vascular endothelium into the blood and the membrane receptor (TCR) for the uptake of TC-Cbl are the two proteins involved in this process. Many aspects of TC are being investigated at this time, however, very little are known about TCR, a protein expressed in actively dividing cells and appear to be cell cycle regulated. We have purified and determined the primary structure of TCR and have identified the gene encoding this protein. Our goal is to understand the genetic regulation of TCR expression and the mechanism by which it binds and mediates the cellular uptake of TC-Cbl to maintain intracellular Cbl. This information will be used in devising strategies to target TCR in cancer therapy, to devise therapeutic and preventive strategies for cellular Cbl deficiency and to study the effects of Cbl deficiency in an animal model. Towards achieving these goals the following specific aims are proposed. 1. To characterize the regulation of TCR gene expression. 2. To determine the synthesis, translocation and functions of TCR in the uptake of TCCbl. 3. To evaluate the effects of functional ablation of TCR at the cellular level and in the whole organism. The rationale for the proposed studies is that understanding how this gene functions will allow us to identify differences in expression or control mechanisms between normal replicating cells and cancer cells and an animal model will allow us to examine the metabolic and structural changes due to Cbl deficiency. We expect these studies to reveal new information on the expression of TCR, whether this pathway could be targeted in cancer therapy and provided a model to understand the metabolic basis for the neuropathological presentation of Cbl deficiency.
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