XIAP controls RIPK2 signaling by preventing its deposition in speck-like structures

The receptor interacting serine/threonine kinase 2 (RIPK2) is essential for linking activation of the pattern recognition receptors NOD1 and NOD2 to cellular signaling events. Recently, it was shown that RIPK2 forms higher order molecular structures in vitro, which were proposed to activate signaling. Here, we demonstrate that RIPK2 forms detergent insoluble complexes in the cytosol of host cells upon infection with invasive enteropathogenic bacteria. Formation of these structures occurred after NF-κB activation and depends on the CARD of NOD1 or NOD2. Complex formation upon activation was dependent on RIPK2 autophosphorylation at Y474 and influenced by phosphorylation at S176. Inhibition of activity of the cIAP protein XIAP induced spontaneous complex formation of RIPK2 but blocked NOD1-dependet NF-κB activation. Using immunoprecipitation, we identified 14-3-3 proteins as novel binding partners of non-activated RIPK2, whereas complexed RIPK2 was bound by the prohibitin proteins Erlin-1 and Erlin-2. Taken together, our work reveals novel roles of XIAP, 14-3-3 and Erlin proteins in the regulation of RIPK2 and expands our knowledge on the function of RIPK2 posttranslational modifications in NOD1/2 signaling.