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Session 4: Immunology Poster 6
Identification of new partners of the protein Nod2 and evaluation of their potential role.
Aurore Lecat 1 , E. Di Valentin 1 , M. Fillet 2 , J. Piette 1 , Sylvie Legrand 1
1. GIGA Research, Unit of Virology and Immunology, B34, University of Liege, Belgium
– aurore.lecat@ulg.ac.be 2. GIGA Research, Unit of Chemical Chemistry, B34,
University of Liege, Belgium
The mammalian innate immune system has evolved to detect pathogens-associated
molecular patterns (PAMPS). Recognition of PAMPS involves membrane-spanning proteins
like the Toll-like receptors (TLR) and cytosolic proteins such as the recently identified NODlike
receptors (NLR). Best studied are mutations in Nod2 that are linked to Crohn’disease
(CD) and Blau syndrome. Nod2 was shown to sense the bacterial peptidoglycan subunit
muramyl-dipeptide (MDP) and to subsequently mediate inflammatory responses in cells by
activating the NF-kB and MAPKS signalling pathways.
This project consists in the identification of new Nod2 partners and in the evaluation
of their role in the Nod2 signalling pathway. We chose a proteomic strategy consisting in the
purification of Nod2-containing complexes in stably Nod2-expressing HEK293 cells after
MDP treatment or infection by Listeria monocytogenes followed by Nod2 partners
identification by mass spectrometry.
Among the candidates, we focused on a Jun N-terminal kinase-binding protein, termed
JNKBP1. This protein is ubiquitously expressed and contains twelve WD40 repeats. We
checked interaction between Nod2 and JNKBP1 and showed that both the N-terminal CARDs
and C-terminal LRRs cooperate in the wild type protein for efficient interaction with
JNKBP1. SiRNA-mediated knockdown of endogenous JNKBP1 significantly upregulated the
phosphorylation of IkB-alpha induced by MDP in stably Nod2-expressing HEK293 cells.
These preliminary results suggest that JNKBP1, through Nod2 interaction, could act as a
suppressor of Nod2-mediated NF-kB activation. In the context of the CD, where loss-offunction
variants of Nod2 have been shown to play a causative role, a suppressor of Nod2
signalling could be of special interest.
We plan to determine the modulator effect of JNKBP1 on Nod2-mediated MAPKs activation
as well as on target gene expression. The molecular mechanism by which JNKBP1
modulates Nod2 signalling could be investigated. All of these experiments should be also
performed in a more relevant cellular model expressing both endogenous proteins such as
intestinal epithelial cells. Finally, we will determine whether JNKBP1 expression is
modulated in pro-inflammatory conditions such as in the mucosa of CD patients.
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