Background: For many human cell types including those of the intestinal epithelium and the immune system, rapid proliferation is essential for normal function. Proliferating cells exhibit increased growth and DNA synthesis which generate nutrient demands. In particular, we have shown that rapidly proliferating lymphocytes increase the uptake of the vitamin biotin five fold, suggesting that cell proliferation generates a substantial increase in biotin demand. This demand might arise from increased synthesis of biotin-dependent carboxylases or from biotinylation of histones (or both). Consistent with this hypothesis are our preliminary studies indicating that biotin deficiency causes reduced proliferation and cell cycle arrest. We hypothesize that biotin has an essential role in cell proliferation in some ways analogous to the role of folate in methylation of DNA regulating the cell cycle. Long-term objectives: To elucidate the role(s) of biotin in the regulation of cell cycle progression and to characterize effects of the cell cycle on cellular biotin homeostasis. Specific hypotheses: (1) At certain phases of the cell cycle, lymphocytes increase expression of the biotin transporter gene leading to increased synthesis of transporter and increase of biotin uptake. (2) In addition to holocarboxylase synthesis, biotin is required in a biotinidase-catalyzed, cell-cycle dependent biotinylation of histones.
Specific aims : (1) To quantitate biotin transporter mRNA and transport activity in lymphocytes at various phases of the cell cycle. (2) To quantitate biotinidase mRNA, cellular biotinidase activity, and biotinylation of histones at various phases of the cell cycle in lymphocytes from healthy adults and, for purposes of comparison, from biotinidase-deficient patients. (3) To investigate the intracellular trafficking of biotin and carboxylase activities. (4) To characterize the effect of extracellular biotin concentration in the culture medium on the proliferation rate of lymphocytes. Benefits: The proposed research will likely lead to novel insights into the mechanisms that generate the increased biotin requirement of proliferating cells. These studies are the initial step to determine whether biotin plays an important role in histone modification and cell cycle regulation. The basic knowledge generated may well be relevant to teratogenesis and immune dysfunction caused by biotin deficiency.
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