Fig. 3

Stx4 and Cdo interact physically in differentiating myoblasts, and the t-SNARE domain of Stx4 mediates the interaction with Cdo. a Lysates of 293T cells transfected with Stx4-myc, Cdo, or control vector were subjected to immunoprecipitation with myc and immunoblotting with Cdo or myc antibodies. b Lysates of C2C12 cells from various differentiation time courses were immunoprecipitated with control IgG or anti-Cdo antibody and immunoblotted with antibodies to Stx4, Cdo, and pan-Cadherin as a loading control. c The schematic representation of the domain structure of Cdo. Cdo consists of five immunoglobulin, three fibronectin type III, a single transmembrane domain, and a 270-amino-acid-long intracellular region. d 293T cells were transiently cotransfected with control or Stx4-myc along with either the full length or three deletion mutants of the Cdo’s cytoplasmic region. Forty-eight hours later, cell lysates were subjected to immunoprecipitation with myc antibody followed by immunoblotting with Cdo antibody. Total lysates served as the expression controls. e The schematic representation depicts the domain structure of Stx4 and the deletion of the specific domain; Δ33-153 (the Syntaxin domain deletion), Δ195-262 (the t-SNARE region deletion), and Δ154-194 (the linker region deletion). f 293T cells were transiently cotransfected with control or Cdo along with the full length or deletion mutants of Stx4, and the lysates were pulled down with S-agarose beads followed by immunoblotting with Cdo or GFP antibodies