FIGURE 6.

Phosphorylation of Osx at Ser-73 and Ser-77 by p38. A, C2C12 cells were co-transfected with constitutively active MKK6 (MKK6EE) and the different Osx constructs as indicated using an empty vector as control. B, 5 μg of GST-Osx wild type or the S73A/S77A mutant (mut) were incubated with activated p38 MAPK (200 ng for each condition) in the presence of γ-[³²P]ATP and visualized by SDS-PAGE and autoradiography (left panel) or visualized by immunoblotting against Osx or phosphoserine (pSer) (right panel). Addition of MKK6-activated recombinant p38 is indicated by an asterisk (p38*). C, wild type (wt) and p38α−/− MEF cells were co-transfected with MKK6EE and Osx constructs as indicated. After 24 h, cells were treated with cycloheximide for the indicated times. Osx was detected by immunoblotting. Quantification of the results of three independent experiments is shown below each panel as mean ± S.E.