The kinase activity of Rip2 determines its stability and consequently Nod1- and Nod2-mediated immune responses.
Rip2 (RICK, CARD3) has been identified as a key effector molecule downstream of the pattern recognition receptors, Nod1 and Nod2; however, its mechanism of action remains to be elucidated. In particular, it is unclear whether its kinase activity is required for signaling or for maintaining protein stability. We have investigated the expression level of different retrovirally expressed kinase-dead Rip2 mutants and the role of Rip2 kinase activity in the signaling events that follow Nod1 and Nod2 stimulation. We show that in primary cells expressing kinase-inactive Rip2, protein levels were severely compromised, and stability could not be reconstituted by the addition of a phospho-mimetic mutation in its autophosphorylation site. Consequently, inflammatory cytokine production in response to Nod1 and Nod2 ligands was abrogated both in vitro and in vivo in the absence of Rip2 kinase activity. Our results highlight the central role that Rip2 kinase activity plays in conferring stability to the protein and thus in the preservation of Nod1- and Nod2-mediated innate immune responses.
Nembrini
Chiara
C
Institute of Integrative Biology, Molecular Biomedicine, CH-8952 Zürich, Switzerland.
Kisielow
Jan
J
Shamshiev
Abdijapar T
AT
Tortola
Luigi
L
Coyle
Anthony J
AJ
Kopf
Manfred
M
Marsland
Benjamin J
BJ
D000119:Acetylmuramyl-Alanyl-Isoglutamine / Q000494:pharmacology; D000276:Adjuvants, Immunologic / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D001854:Bone Marrow Cells / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000097:blood / Q000378:metabolism; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D055879:Gene Knock-In Techniques; D005816:Genetic Complementation Test; D018836:Inflammation Mediators / Q000097:blood / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D053474:Nod1 Signaling Adaptor Protein / Q000235:genetics / Q000378:metabolism*; D053473:Nod2 Signaling Adaptor Protein / Q000235:genetics / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012772:Shock, Septic / Q000097:blood / Q000276:immunology
D000276:Adjuvants, Immunologic; D016207:Cytokines; D018836:Inflammation Mediators; D008070:Lipopolysaccharides; D053474:Nod1 Signaling Adaptor Protein; C507873:Nod1 protein, mouse; D053473:Nod2 Signaling Adaptor Protein; C495945:Nod2 protein, mouse; D009842:Oligopeptides; D000119:Acetylmuramyl-Alanyl-Isoglutamine; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C506255:Ripk2 protein, mouse; C039537:heptanoyl-gamma-D-glutamyl-L-meso-diaminopimelyl-D-alanine