FIGURE 5.

Osx phosphorylation by p38. A, MC3T3-E1 cells were treated with or without BMP-2 in medium without serum. C2C12 cells were transfected with empty vector, short Osx, or long Osx constructs. After 24 h, cell extracts were incubated with alkaline phosphatase (ALP) for 1 h. Osx forms were detected by immunoblotting. B, cells were treated with or without BMP-2 in medium without serum for 24 h and pretreated with SB203580 (SB) for 30 min where indicated. Osx and tubulin were detected by immunoblotting. C, C2C12 cells were co-transfected with constitutively active MKK6 (MKK6EE) and short or long Osx constructs. After 24 h, cells were treated with SB203580 for 3 h. Osx, MKK6, and tubulin were detected by immunoblotting. D, wild type (wt) and p38α−/− MEF cells were co-transfected with MKK6 construct and the short Osx construct. After 24 h, cells were treated with SB203580 for 3 h. Osx, MKK6, and tubulin were detected by immunoblotting. Arrows indicate phosphorylated forms.