Cloning and Characterization of Mouse Deoxyguanosine Kinase

Abstract

Deoxyguanosine kinase (dGK) is a nuclear gene product that catalyzes the phosphorylation of purine deoxyribonucleosides and their analogues. The human enzyme is located predominantly in the mitochondria, as shown by biochemical fractionation studies and in situ localization of the overexpressed recombinant protein.Here we describe the cloning of mouse dGK cDNA and the identification of a novel amino-terminally truncated isoform that corresponds to about 14% of the total dGK mRNA population in mouse spleen. In situ fluorescence assays suggest that the new isoform cannot translocate into the mitochondria and thus may represent a cytoplasmic enzyme. Expression of mouse dGK mRNA was highly tissue-specific and differed from the tissue distribution observed in humans. Recombinant mouse dGK showed similar specific activity and substrate specificity as compared with the human enzyme. The broad specificity, restricted tissue distribution, and location of mouse dGK in multiple cellular compartments raise new considerations with respect to the role of the individual deoxynucleoside kinases in nucleotide metabolism.

Apoptosis Induces Efflux of the Mitochondrial Matrix Enzyme Deoxyguanosine Kinase

“…4A). The punctuate labeling pattern around the nuclei was identical to that previously reported for HEK 293 cells (13) and similar to that obtained when a mouse dGK was fused to green fluorescent protein and overexpressed in Cos-1 cells (23). The majority of the CdCl 2 -treated cells showed a diffuse cytosolic staining pattern with the dGK antibody (Fig.…”

“…Six h after addition of CdCl 2 to HEK 293 cells, dGK was localized extra-mitochondrially in many cells. The localization pattern was similar to that reported for the cytosolic isoform of mouse dGK (23) and Smac/DIABLO 4 h after induction of apoptosis (24). In many CdCl 2 -treated cells, the overall dGK labeling was more intense compared with the control cells.…”

Nucleoside analogues: mechanisms of drug resistance and reversal strategies

“…[133][134][135][136] Cytoplasmic dGK may also contribute to the supply of purine deoxynucleotides for nuclear DNA replication and repair. 132 In this way, nucleoside ana- logues phosphorylated by dGK may exert their cytotoxic effects by interference with both mitochondrial and nuclear DNA synthesis. Whether inactivation of dGK is involved in resistance to NA is not presently known.…”

Mitochondrial and submitochondrial localization of human deoxyguanosine kinase

“…A potential cytosolic isoform of deoxyguanosine kinase was recently reported in mouse cells [11], and therefore we also used the deoxyguanosine kinase antibody to study the in situ distribution of this enzyme in human embryonic kidney (HEK) 293 cells by immunohistochemistry. …”