title: CK2 phosphorylation of human centrins 1 and 2 regulates their binding to the DNA repair protein XPC, the centrosomal protein Sfi1 and the phototransduction protein transducin β.
abstract: Centrins are calcium-binding proteins that can interact with several cellular targets (Sfi1, XPC, Sac3 and transducin β) through the same hydrophobic triad. However, two different orientations of the centrin-binding motif have been observed: W(1)xxL(4)xxxL(8) for XPC (xeroderma pigmentosum group C protein) and the opposite orientation L(8)xxxL(4)xxW(1) for Sfi1 (suppressor of fermentation-induced loss of stress resistance protein 1), Sac3 and transducin β. Centrins are also phosphorylated by several protein kinases, among which is CK2. The purpose of this study was to determine the binding mechanism of human centrins to three targets (transducin β, Sfi1 and XPC), and the effects of in vitro phosphorylation by CK2 of centrins 1 and 2 with regard to this binding mechanism. We identified the centrin-binding motif at the COOH extremity of transducin β. Human centrin 1 binds to transducin β only in the presence of calcium with a binding constant lower than the binding constant observed for Sfi1 and for XPC. The affinity constants of centrin 1 were 0.10 10(6) M(-1), 249 10(6) M(-1) and 52.5 10(6) M(-1) for Trd, R17-Sfi1 and P17-XPC respectively. CK2 phosphorylates human centrin 1 at residue T138 and human centrin 2 at residues T138 and S158. Consequently CK2 phosphorylation abolished the binding of centrin 1 to transducin β and reduced the binding to Sfi1 and XPC. CK2 phosphorylation of centrin 2 at T138 and S158 abolished the binding to Sfi1 as assessed using a C-HsCen2 T138D-S158D phosphomimetic form of centrin 2.
authors.affiliation: Institut Curie-Centre de Recherche, F-91405 Orsay Cédex, France.
authors.forename: Dora
authors.initials: D
authors.lastname: Grecu
authors.affiliation: Institut Curie-Centre de Recherche, F-91405 Orsay Cédex, France.
authors.forename: Liliane
authors.initials: L
authors.lastname: Assairi
keywords: C-HsCen, C-terminal domain of centrin; CK2; CK2, casein protein kinase; Centrins; HPLC, high pressure liquid chromatography; HsCen, human centrin; ITC, isothermal titration calorimetry; MALDI-TOF, matrix-assisted laser desorption ionization time of flight; Sfi1; Sfi1, suppressor of fermentation-induced loss of stress resistance protein 1; Transducin; Trd, transducin; XPC; XPC, xeroderma pigmentosum group C protein