Identification of a putative human mitochondrial thymidine monophosphate kinase associated with monocytic/macrophage terminal differentiation.
Mitochondrial DNA synthesis requires the supply of thymidine triphosphate (dTTP) independent of nuclear DNA replication. In resting and differentiating cells that withdraw from the cell cycle, mitochondrial thymidine kinase 2 (TK2) mediates thymidine monophosphate (dTMP) formation for the dTTP biosynthesis in mitochondria. However, a thymidine monophosphate kinase (TMPK) that phosphorylates dTMP to form thymidine diphosphate (dTDP) in mitochondria remains undefined. Here, we identified an expressed sequence tag cDNA, which encodes a TMPK with a mitochondrial import sequence at its N-terminus designated as TMPK2. HeLa cells expressing TMPK2 fused to green fluorescent protein (GFP) displayed green fluorescence in mitochondria. Over-expression of TMPK2 increased the steady-state level of cellular dTTP and promoted the conversion of radioactive labeled-thymidine and -dTMP to dTDP and dTTP in mitochondria. TMPK2 RNA was detected in several tissues and erythroblastoma cell lines. We also generated TMPK2 antibody and used it for immunofluorescence staining to demonstrate endogenous expression of TMPK2 in mitochondria of erythroblastoma cells. Finally, we showed that TMPK2 protein expression was upregulated in monocyte/macrophage differentiating cells, suggesting the coordinated regulation of TMPK2 expression with the terminal differentiation program.
Chen
Yen-Ling
YL
Graduate Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, No. 1, Section 1, Jen-Ai Road, Taipei 100, Taiwan.
Lin
Da-Wei
DW
Chang
Zee-Fen
ZF
D000595:Amino Acid Sequence; D002454:Cell Differentiation / Q000502:physiology*; D002460:Cell Line; D003001:Cloning, Molecular; D006367:HeLa Cells; D006801:Humans; D008264:Macrophages / Q000166:cytology* / Q000201:enzymology; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D009000:Monocytes / Q000166:cytology* / Q000201:enzymology; D009703:Nucleoside-Phosphate Kinase / Q000235:genetics / Q000502:physiology*; D013942:Thymine Nucleotides / Q000096:biosynthesis
D013942:Thymine Nucleotides; D009703:Nucleoside-Phosphate Kinase; C020093:dTMP kinase; C024157:thymidine 5'-triphosphate