Csapo Zsolt, Keszler Gergely, Safrany Geza, Spasokoukotskaja Tatjana, Talianidis Iannis, Staub Maria, Sasvari-Szekely Maria
Institute of Medical Chemistry, Molecular Biology and Pathobiochemistry, Semmelweis University, P.O. Box 260, H-1444 Budapest, Hungary.
Biochem Pharmacol. 2003 Jun 15;65(12):2031-9. doi: 10.1016/s0006-2952(03)00182-5.
Deoxycytidine kinase (dCK) is a key enzyme in the intracellular metabolism of deoxynucleosides and their analogues, phosphorylating a wide range of drugs used in the chemotherapy of leukaemia and solid tumours. Previously, we found that activity of dCK can be enhanced by incubating primary cultures of lymphocytes with substrate analogues of the enzyme, as well as with various genotoxic agents. Here we present evidence that exposure of human lymphocytes to 0.5-2 Gy dosage of gamma-radiation as well as incubation of cells with calyculin A, a potent inhibitor of protein phosphatase 1 and 2A, both elevate dCK activity without changing the level of dCK protein. When cells were gamma-irradiated in the presence of calyculin A, a more pronounced activation of dCK was observed. In contrast, both basal and stimulated dCK activities were reduced by hyperosmotic treatment of the cells. DNA repair determined by the Comet assay and by thymidine incorporation was induced by irradiation. Complete repair of gamma-irradiated DNA was detected within 1 hr following the irradiation along with dCK activation, but the rate of repair was not accelerated by calyculin A. These data provide evidence for the activation of dCK upon DNA damage and repair that seems to be mediated by phosphorylation of the enzyme, suggesting the role of dCK in DNA repair processes.
脱氧胞苷激酶（dCK）是脱氧核苷及其类似物细胞内代谢中的关键酶，可磷酸化用于白血病和实体瘤化疗的多种药物。此前，我们发现，通过用该酶的底物类似物以及各种基因毒性剂孵育淋巴细胞原代培养物，dCK的活性可以增强。在此，我们提供证据表明，人淋巴细胞暴露于0.5 - 2 Gy剂量的γ辐射以及用花萼海绵诱癌素A（一种蛋白磷酸酶1和2A的强效抑制剂）孵育细胞，均可提高dCK活性，而不改变dCK蛋白水平。当细胞在花萼海绵诱癌素A存在下进行γ辐射时，观察到dCK有更明显的激活。相反，细胞的高渗处理会降低基础和刺激后的dCK活性。通过彗星试验和胸苷掺入法测定的DNA修复由辐射诱导。在辐射后1小时内检测到γ辐射DNA的完全修复以及dCK激活，但花萼海绵诱癌素A并未加速修复速率。这些数据为DNA损伤和修复时dCK的激活提供了证据，这种激活似乎是由该酶的磷酸化介导的，提示dCK在DNA修复过程中的作用。
