Role for erbin in bacterial activation of Nod2.
Intracellular peptidoglycan (PG) recognition in human cells is mediated by the NACHT-LRR proteins Nod1 and Nod2. Elicitation of these proteins by PG motifs released from invasive bacteria triggers signaling events, resulting in the activation of the NF-kappaB pathway. In order to decipher the molecular components involved in Nod2 signal transduction, we set out to identify new interaction partners of Nod2 by using a yeast two-hybrid screen. Besides the known interaction partner RIP2, the screen identified the leucine-rich repeat (LRR)- and PDZ domain-containing family member Erbin as a binding partner of Nod2. Erbin showed a specific interaction with Nod2 in coimmunoprecipitation experiments with human HEK 293T cells. Immunofluorescence microscopy with a newly generated anti-Nod2 monoclonal antibody showed that Erbin and Nod2 partially colocalize in human cells. Subsequent analysis of the Erbin/Nod2 interaction revealed that the LRR of Erbin and the caspase activating and recruiting domains of Nod2 were necessary for this interaction. No significant interaction was observed with a Walker B box mutant of Nod2 or a Crohn's disease-associated frameshift mutant of Nod2, indicating that complex formation is dependent on the activity of the molecule. In addition, a change in the dynamics of the Erbin/Nod2 complex was observed during Shigella flexneri infection. Furthermore, ectopic expression of increasing amounts of Erbin or short hairpin RNA-mediated knockdown of Erbin showed a negative influence of Erbin on Nod2/muramyl-dipeptide-mediated NF-kappaB activation. These results implicate Erbin as a potential negative regulator of Nod2 and show that bacterial infection has an impact on Nod2/Erbin complex formation within cells.
Kufer
T A
TA
Immunité Innée et Signalisation, Institut Pasteur, 28 rue du Docteur Roux, 75724 Paris, France.
Kremmer
E
E
Banks
D J
DJ
Philpott
D J
DJ
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000502:physiology*; D002478:Cells, Cultured; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000502:physiology*; D016328:NF-kappa B / Q000378:metabolism; D053473:Nod2 Signaling Adaptor Protein; D017434:Protein Structure, Tertiary; D012763:Shigella flexneri / Q000472:pathogenicity*
D048868:Adaptor Proteins, Signal Transducing; C413364:ERBIN protein, human; D047908:Intracellular Signaling Peptides and Proteins; D016328:NF-kappa B; C429018:NOD2 protein, human; D053473:Nod2 Signaling Adaptor Protein