Figure 5

CK2 phosphorylates C1ORF109 in vitro . (A) C1ORF109-Flag was immunoprecipitated from HEK293 cells and was immunoblotted for anti-phosphoserine. (B) C1ORF109-Flag purified from HEK293 cells was treated or mock treated with calf intestinal phosphatase (CIP) and immunoblotted for C1ORF109. (C) C1ORF109-Flag immunoprecipitated from HEK293 cells was incubated with human CK2, and [γ-³²P]-ATP, and the CPM was read in a scintillation counter subsequently. (D) C1ORF109-Flag purified from HEK293 cells was phosphorylated with human CK2 and [γ-³²P]-ATP, fractionated by SDS-PAGE, and dried Coomassie blue-stained gels were examined by autoradiography. Heparin was used at 10 μg/ml. (E) Wild type C1ORF109 and C1ORF109Mut immunoprecipitated from HEK293 cells were incubated with human CK2, and [γ-³²P]-ATP respectively, and the CPM was read in a scintillation counter subsequently.