In the process of identifying genes involved in optimization of lymphocyte activation, we have cloned the human mitochondrial NADP-dependent isocitrate dehydrogenase (mNADP-IDH) cDNA. The cDNA and its deduced amino acid (AA) sequence had a high degree of homology with those of the porcine and bovine. The heart and muscle had the highest constitutive expression of the gene. The expression of steady-state mRNA in the resting T and B lymphocytes was low but was induced after mitogen stimulation. The mRNA levels peaked around 48 h and remained elevated at 72 h. At the protein level, the mitochondrial but not cytosolic NADP-IDH activity was augmented after the mitogen stimulation. There was no cell cycle-dependent fluctuation of mNADP-IDH expression in synchronized Jurkat cells. In T and B cells, rapamycin (RAPA) could repress the mitogen-stimulated mNADP-IDH expression, although most of the early or late phase activation-related genes including a G-protein beta subunit-related gene H12.3 were not affected by the drug. The restricted expression of the gene in certain tissues and the activation-related expression in lymphocytes suggest that this gene might be necessary for optimal functions in heart, muscle, and the activated lymphocytes.