Biochemical and Biophysical Research Communications
Nod1, a CARD protein, enhances pro-interleukin-1β processing through the interaction with pro-caspase-1☆
Section snippets
Materials and methods
Plasmids. The expression plasmids, pcDNA3-Myc-caspase-1 (wild type), pcDNA3-caspase-1-Myc (wild type), pcDNA3-Myc-caspase-1 (Cys 285 Ser), pcDNA3-HA-caspase-1 (Cys 285 Ser), pcDNA3-Flag-RIP2, and pCMV-IL-1β, have been described previously [21]. pcDNA3-Nod1-Flag, pcDNA3-Flag-Nod1-CARD (residues 1–158), pcDNA3-Myc-RIP2-CARD (residues 365–531), pcDNA3-Flag-ICEBERG, and pcDNA3-Myc-Nod1 were created by polymerase chain reaction (PCR).
Co-immunoprecipitations and immunoblotting assays. Human embryonic
Nod1 binds to both caspase-1 and RIP2 through CARD–CARD interaction
Because an earlier report described that Nod1 interacts both with pro-caspase-1 and RIP2 [13], we tested the possibility that these interactions are mediated through heterotypic CARD–CARD interactions. For these experiments, 293T cells were transiently transfected with expression plasmids encoding Flag-tagged-Nod1-CARD either with an expression plasmid producing HA-tagged caspase-1-CARD or Myc-tagged RIP2-CARD protein. Immunoprecipitations were then performed with anti-Flag antibody and the
Discussion
It has been known that pro-caspase-1 interacts with Nod1 [13]. We therefore undertook a study of the consequences of this interaction. Because the key role of caspase-1 is IL-1β processing, we investigated whether Nod1 influences IL-1β secretion induced by the expression of caspase-1. Nod1 interacts with caspase-1 through CARD–CARD interactions, thereby promoting caspase-1 oligomerization (Fig. 1). Consequently, Nod1 expression increases caspase-1-induced secretion of mature IL-1β in 293T
Acknowledgements
This work was supported by the KOSEF through the Cell death Disease Center at the Catholic University of Korea.
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2014, Trends in ImmunologyCitation Excerpt :The guanosine nucleotide-exchange factor activity of SopE is required for NF-κB activation [75] and for the recruitment of NOD1 to membrane ruffles [40]. The observation that SopE can activate inflammasome-dependent cytokine maturation [76,77] might be a consequence of nodosome activation, because NOD1 is known to enhance interleukin (IL)-1β secretion [78]. IL-1β secretion requires two signals.
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Abbreviations: IL-1β, interleukin-1β; CARD, caspase recruit domain; NOD, nucleotide-binding oligomerization domain; PCR, polymerase chain reaction; DAPI, 4′,6-diamidine-2′-phenylindole dihydrochloride; GFP, green fluorescent protein; SDS, sodium dodecyl sulfate; PAGE, polyacrylamide gel electrophoresis; LPS, lipopolysaccharide; LRR, leucine-rich repeat.