Casein kinase 1delta activates human recombinant deoxycytidine kinase by Ser-74 phosphorylation, but is not involved in the in vivo regulation of its activity.
Deoxycytidine kinase (dCK) is a key enzyme in the salvage of deoxynucleosides and in the activation of several anticancer and antiviral nucleoside analogues. We recently showed that dCK was activated in vivo by phosphorylation of Ser-74. However, the protein kinase responsible was not identified. Ser-74 is located downstream a Glu-rich region, presenting similarity with the consensus phosphorylation motif of casein kinase 1 (CKI), and particularly of CKI delta. We showed that recombinant CKI delta phosphorylated several residues of bacterially overexpressed dCK: Ser-74, but also Ser-11, Ser-15, and Thr-72. Phosphorylation of dCK by CKI delta correlated with increased activity reaching at least 4-fold. Site-directed mutagenesis demonstrated that only Ser-74 phosphorylation was involved in dCK activation by CKI delta, strengthening the key role of this residue in the control of dCK activity. However, neither CKI delta inhibitors nor CKI delta siRNA-mediated knock-down modified Ser-74 phosphorylation or dCK activity in cultured cells. Moreover, these approaches did not prevent dCK activation induced by treatments enhancing Ser-74 phosphorylation. Taken together, the data preclude a role of CKI delta in the regulation of dCK activity in vivo. Nevertheless, phosphorylation of dCK by CKI delta could be a useful tool for elucidating the influence of Ser-74 phosphorylation on the structure-activity relationships in the enzyme.
Smal
Caroline
C
Laboratory of Physiological Chemistry, Université catholique de Louvain, de Duve Institute, B-1200 Brussels, Belgium. caroline.smal@uclouvain.be
Vertommen
Didier
D
Amsailale
Rachid
R
Arts
Angélique
A
Degand
Hervé
H
Morsomme
Pierre
P
Rider
Mark H
MH
Neste
Eric Van Den
EV
Bontemps
Françoise
F
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D048148:Casein Kinase Idelta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D003842:Deoxycytidine Kinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011499:Protein Processing, Post-Translational; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000737:chemistry
D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D012694:Serine; D003842:Deoxycytidine Kinase; D048148:Casein Kinase Idelta