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. 2013 Sep 15;24(18):2907–2917. doi: 10.1091/mbc.E13-01-0014

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© 2013 Arasaki et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 4: — Interaction of RINT-1 with Cog1. (A, B) Solubilized membrane fractions of 293T cells were incubated for 1 h with an antibody against RINT-1 (lane 2) or ZW10 (lane 3). Immunocomplexes were precipitated with protein G–Sepharose, subjected to SDS–PAGE, and then analyzed by immunoblotting with the indicated antibodies. Three percent input was also analyzed (lane 1). (C) Each of the FLAG-Cog subunits was coexpressed with GFP–RINT-1 in 293T cells. Cell lysates were immunoprecipitated with anti-FLAG beads, subjected to SDS–PAGE, and then analyzed by immunoblotting with antibodies against GFP (middle) and FLAG (bottom). Five percent input was also analyzed by immunoblotting with an anti-GFP antibody (top). (D) Each of the GFP-tagged RINT-1 constructs was coexpressed with FLAG-Cog1, and cell lysates were immunoprecipitated with anti-FLAG beads, subjected to SDS–PAGE, and analyzed by immunoblotting with antibodies against GFP (middle) and FLAG (bottom). Five percent input was also analyzed by immunoblotting with an anti-GFP antibody (top). (E) GST fused to each of the Cog1 constructs was coexpressed with FLAG–RINT-1, and cell lysates were pulled down with glutathione beads and analyzed by immunoblotting with antibodies against FLAG (middle) and GST (bottom). Five percent input was also analyzed by immunoblotting with an anti-FLAG antibody (top).