FIGURE 7.

JNKBP1 interferes with NOD2 oligomerization and NOD2-mediated RIP2 tyrosine phosphorylation. A, JNKBP1 interferes with NOD2 oligomerization. GFP-NOD2 and HA-NOD2 were transfected in HEK293T cells with an empty or JNKBP1-V5 expression vector. After 48 h, cells were treated with MDP (10 μg/ml) for 30 min before being harvested for protein extraction. Samples were immunoprecipitated with an anti-HA antibody. The self-association of NOD2 and the presence of JNKBP1 in the NOD2-containing complex were analyzed by anti-GFP and anti-V5 immunoblot, respectively. Total lysates were immunoblotted with anti-GFP, -HA, and -V5 antibodies. B, JNKBP1 interferes with NOD2-mediated RIP2 tyrosine phosphorylation. FLAG-RIP2 and GFP-NOD2 were transfected in HEK293T cells with an empty or JNKBP1 expression vector. After 48 h, cells were harvested for protein extraction. Proteins were immunoprecipitated with an anti-FLAG antibody. The RIP2 tyrosine phosphorylation and the presence of NOD2 in the RIP2-containing complex were analyzed by anti-phosphotyrosine and anti-GFP immunoblots, respectively. Total lysates were immunoblotted with anti-GFP, -FLAG, and -V5 antibodies.