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. 2009 May 26;29(15):4130–4143. doi: 10.1128/MCB.00199-09

FIG. 3.

FIG. 3.

The Abl-binding motif of Cdo is functional in myogenic differentiation. (A) Photomicrographs of C2C12 cells that stably express the indicated Cdo proteins or a control vector (pBp) that were cultured in DM, fixed, and stained with an antibody to MHC. (B) Quantification of myotube formation by the cell lines shown in panel A. Values represent means of triplicate determinations ± 1 standard deviation. The experiment was repeated three times with similar results. Asterisks indicate difference from the control at P < 0.01. (C) Lysates of cell lines shown in panel A were Western blotted with antibodies to Cdo, MHC, and β-tubulin as a loading control. (D) Photomicrographs of C2C12 cells that stably express the indicated Cdo proteins or a control vector (pBp) that were cultured in DM, fixed, and stained with an antibody to MHC. (E) Quantification of myotube formation by the cell lines shown in panel D. Values represent means of triplicate determinations ± 1 standard deviation. The experiment was repeated three times with similar results. Asterisks indicate difference from the control at P < 0.01. (F) Lysates of cell lines shown in panel D were Western blotted with antibodies to Cdo and pan-cadherin as a loading control. (G) Lysates of cell lines shown in panel D were cultured for 2 or 3 days in DM (D2 and D3, respectively) and then Western blotted with antibodies to MHC and pan-cadherin as a loading control. (H) C2C12 cells were cotransfected with control or Cdo siRNA expression vectors plus Cdo or CdoP/A expression vectors and pQ-lacZ (a vector that drives expression of nucleus-localized β-Gal to mark transfectants). Differentiated cultures were double stained for β-Gal activity (blue color) and for MHC expression (brown color). (I) Quantification of C2C12 cell differentiation shown in panel H. Cultures were scored as MHC− or MHC+, with MHC+ cells further scored as having a single nucleus, two to four nuclei, or five or more nuclei. Values represent means of triplicate determinations ± 1 standard deviation. The experiment was repeated three times with similar results.