Figure 6.

CK2β enhances TGFβ1- and BMP-9 stimulated Smad1/5/8 phosphorylation. A) P19 cells were transfected with ALK-1, CK2β expression vector, or siRNA to CK2β, as indicated. Twenty-four hours after transfection, cells were serum starved for 12 h and then stimulated with the indicated amounts of TGFβ1 or BMP-9 for the indicated periods of times, the cells were lysed, and the lysates were assayed for pSmad1 and total Smad1 expression by immunoblot analysis. Portions of the lysates were analyzed by immunoblot for the indicated proteins to confirm expression and/or knockdown of CK2β expression. Data are representative of 3 independent experiments. B, C) Quantification of Smad-1 phosphorylation was performed with the assistance of the ImageJ analysis program. Integrated density measurements for each variable were taken, and background was subtracted. A ratio of phosphorylated/total Smad-1 was then calculated for every condition, and this was then normalized against the ALK-1 signal obtained for that condition.