Figure 6

VP2 binds to chromatin and interacts with MCM3 but this does not require dual phosphatase activity. (A) The soluble (S) and chromatin (C) fractions were prepared by using CSK buffer containing 0.5% Triton X-100 in CHO cells of transfected with VP2-GFP plasmid. The fractions were treated with 0 U (-) and 150 U (+) of MNase or the indicated concentrations of NaCl. Nuclear extracts (N) containing VP2-GFP were as positive control and all fractions were subjected to immunoblotting against with lamin B receptor, MCM3, and VP2 antibodies. (B) At 48 h post-transfection, the GFP (as Control) and Flag-VP2-GFP (as wild-type; WT) were found in the CHO cells. Next the lysates were immunoprecipitated using Flag M2 beads and immunoblotted against VP2 and MCM3 antibodies. (C) The WT and mutants (C95S, C97S, and C95S/C97S) of the Flag-VP2-GFP in CHO cells were also examined at 48 h post-transfection. The cell lysates were immunoprecipitated using Flag M2 beads and immunoblotted against with VP2 and MCM3 antibodies. The arrow head was indicated a none-specific band. The nuclear extracts containing VP2-GFP were designated as N