2. Regulation of deoxycytidine kinase activity
Deoxycytidine kinase (dCK) catalyzes the phosphorylation of deoxycytidine, deoxyadenosine and deoxyguanosine, which is the rate-limiting step in the deoxynucleoside salvage pathway. Moreover, dCK phosphorylates and activates numerous nucleoside analogues (NA) used against leukaemias, solid tumors and viral infections. The enzyme is preferentially expressed in lymphoid cells, which explains the clinical success of NA against lymphoproliferative disorders, such as hairy cell leukaemia and B-CLL. Because dCK plays an essential role in the therapeutic efficacy of NA, identification of mechanisms that control dCK activity are of particular interest.
In the recent years, dCK was found to be activated by several genotoxic agents. It was suggested that this activation could result from a phosphorylation process. Using HEK (human embryonic kidney) 293 cells overexpressing dCK, we have been the first to demonstrate that dCK is a phosphoprotein and that dCK activity is strongly correlated to phosphorylation of Ser-74 (Smal et al., J. Biol. Chem. 2006). We are now attempting to identify the protein kinase(s) responsible for the phosphorylation of dCK on Ser-74 and the signalling pathway that leads to dCK activation following treatment with DNA damaging agents.
Three-dimensional structure of deoxycytidine kinase in complex with ADP and deoxycytidine
The Ser-74 phosphorylation site is located at a very flexible 15-residue insert (Ser-63 - Asn-77) easily accessible to cellular protein kinases.
Using an anti-phospho-Ser-74 antibody, we have also showed that endogenous dCK is phosphorylated on Ser-74 in B-CLL lymphocytes and that genotoxic agents increase Ser-74 phosphorylation, in close parallel with changes in dCK activity (Smal et al., Cancer Lett. 2007). These results suggest that activation of dCK via phosphorylation of Ser-74 might constitute a new therapeutic strategy to enhance efficacy of NA. Studies are in progress to assess the relationship between phosphorylation of dCK on Ser-74, activation of various NA, and in vitro sensitivity to the latter drugs.
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