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. 2012 Apr 23;287(24):20081–20087. doi: 10.1074/jbc.M111.335463

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© 2012 by The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 1. — Identification of PKCδ as a CARMA1-associated protein. A, interaction of overexpressed HA-PKCδ and Flag-CARMA1. Expression plasmids encoding HA-PKCδ (10 μg) and Flag-CARMA1 (4 μg) were transfected into HEK293 cells. Lysates were immunoprecipitated with antibodies to Flag (left panel), HA (right panel), or control IgG, and then analyzed by immunoblotting with an antibody to HA (upper left panel) or Flag (upper right panel). Expression of the transfected proteins were analyzed by immunoblotting with antibodies to Flag and HA (lower panel). B, PKCδ interacts with MALT1 and TRAF6 but not Bcl10. HEK293 cells were transfected with the indicated plasmids. Coimmunoprecipitation and immunoblot analysis were performed with the indicated antibodies. C and D, CD3/CD28 costimulation induces the association between PKCδ and CARMA1. Jurkat T cells were stimulated by CD3/CD28 crosslinking for the indicated time. Lysates were immunoprecipitated with an antibody to CARMA1 (C) or PKCδ (D). The precipitates and lysates were analyzed by immunoblotting with the indicated antibodies. Ig, control mouse IgG; αF, anti-Flag; Pre, preimmune serum; *, nonspecific band.