ABSTRACT
Nucleotide-binding oligomerization domain protein 1 (NOD1) belongs to a family that includes multiple members with NOD and leucine-rich repeats in vertebrates and plants. NOD1 has been suggested to have a role in innate immune responses, but the mechanism involved remains unknown. Here we report that NOD1 mediates the recognition of peptidoglycan derived primarily from Gram-negative bacteria. Biochemical and functional analyses using highly purified and synthetic compounds indicate that the core structure recognized by NOD1 is a dipeptide, gamma-D-glutamyl-meso-diaminopimelic acid (iE-DAP). Murine macrophages deficient in NOD1 did not secrete cytokines in response to synthetic iE-DAP and did not prime the lipopolysaccharide response. Thus, NOD1 mediates selective recognition of bacteria through detection of iE-DAP-containing peptidoglycan.
Subject(s)
Adaptor Proteins, Signal Transducing , Bacterial Infections/immunology , Carrier Proteins/physiology , Diaminopimelic Acid/metabolism , Immunity, Innate/physiology , Intracellular Signaling Peptides and Proteins , Peptidoglycan/metabolism , Animals , Cell Line , Cytokines/biosynthesis , Female , Humans , Lipopolysaccharides/metabolism , Mice , Mice, Inbred C57BL , Nod1 Signaling Adaptor Protein , Nod2 Signaling Adaptor ProteinABSTRACT
Genetic variation in human Nod2 has been associated with susceptibility to Crohn's disease. The mouse Nod2 locus is located at chromosome 8 and composed of 12 exons, 11 of which encode the Nod2 protein. Sequence analysis of Nod2 from 45 different strains of Mus musculus and Mus spretus revealed extensive polymorphism involving all exons of Nod2. Of the 140 polymorphic sites identified, 68 were located in the coding region, of which 28 created amino acid substitutions in Nod2. Expression of mouse Nod2 activated NF-kappaB and conferred responsiveness to bacterial components, an activity that was deficient in mutants corresponding to those associated with susceptibility to Crohn's disease. These studies demonstrate a conserved role for Nod2 in the response to bacterial components and suggest that selective evolutionary pressure exerted by pathogens may have contributed to the high level of variability of Nod2 sequences in both humans and mice.
Subject(s)
Carrier Proteins/genetics , Crohn Disease/genetics , Genetic Variation/genetics , Intracellular Signaling Peptides and Proteins , Mice/genetics , Muridae/genetics , Amino Acid Sequence , Animals , Chromosome Mapping , DNA Primers , Gene Expression , Humans , Molecular Sequence Data , Nod2 Signaling Adaptor Protein , Phylogeny , Polymorphism, GeneticABSTRACT
BACKGROUND & AIMS: The NOD2 variants R702W, G908R, and L1007fsinsC are strongly associated with Crohn's disease (CD) in both European and American populations, but whether this susceptibility extends to all ethnic groups remains unknown. Except for the L1007fsinsC mutation, which produces a truncated NOD2 protein, the functional activity of the major CD-associated variants G908R and R702W is unknown. METHODS: Individuals were genotyped for R702W, G908R, and L1007fsinsC. The ability of G908R, R702W, and L1007fsinsC variants in the presence and absence of P268S to confer responsiveness to lipopolysaccharide (LPS) and peptidoglycan (PGN) was determined in HEK293T kidney cells. RESULTS: G908R and L1007fsinsC, but not R702W, were associated with disease susceptibility in Ashkenazi Jews. Ashkenazi Jews with CD had significantly higher allele frequency carriage of G908R and lower carriage of R702W compared with non-Jewish whites with CD. Functional studies revealed that the G908R, R702W, and L1007fsinsC variants in the presence and absence of P268S are defective in their ability to respond to bacterial LPS and PGN, whereas P268S alone exhibited wild-type activity. CONCLUSIONS: R702W is not associated with susceptibility to CD in Ashkenazi Jews. The G908R, R702W, and L1007fsinsC variants share a common signaling defect in response to bacterial components, providing evidence for a unifying molecular mechanism whereby NOD2 mutations contribute to disease susceptibility.
