Molecular cloning of the cDNA of mouse mitochondrial NADP-dependent isocitrate dehydrogenase and the expression of the gene during lymphocyte activation.
The current report documents the molecular cloning of the mouse mitochondrial NADP-dependent isocitrate dehydrogenase (mNADP-IDH) cDNA. The cDNA was 1,863 bp in length and contained one open reading frame encoding a 523-residue polypeptide with a predicted molecular weight of 58 kDa. The cDNA and the deduced amino acid (AA) sequence of the mouse mNADP-IDH had a high degree of homology with those of porcine, bovine, alfalfa, and yeast. The recombinant mNADP-IDH expressed in Escherichia coli had active enzymatic function, as well as an expected molecular weight. The heart had the highest constitutive expression of the steady-state mNADP-IDH mRNA, followed by the kidney, while the expression of the gene in other tissues was low. The enzymatic activity of different tissues was in agreement with their mNADP-IDH mRNA levels. The resting lymphocytes had low constitutive expression of the gene, but the steady-state mRNA could be induced 48 h after mitogen stimulation. At the protein level, the resting lymphocytes had low enzymatic activity of mNADP-IDH, but the activity was augmented fivefold after mitogen stimulation. The cytosolic NADP-IDH, on the contrary, remained low or undetectable before and after the mitogen stimulation. Based on our current findings as well as the known roles of the mNADP-IDH in anabolism and in the isocitrate shuttle, it is conceivable that the mNADP-IDH is necessary for optimizing proliferation in lymphocytes.
Yang
L
L
Department of Medicine, Notre-Dame Hospital Research Center and Nephrology Service, Montreal, Québec, Canada.
Luo
H
H
Vinay
P
P
Wu
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D016193:G1 Phase / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000276:immunology* / Q000502:physiology; D015723:Gene Library; D006801:Humans; D007521:Isocitrate Dehydrogenase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008213:Lymphocyte Activation / Q000235:genetics*; D008214:Lymphocytes / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009928:Organ Specificity; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid
D018076:DNA, Complementary; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D007521:Isocitrate Dehydrogenase; C039355:isocitrate dehydrogenase (NADP+)