A novel PAAD-containing protein that modulates NF-kappa B induction by cytokines tumor necrosis factor-alpha and interleukin-1beta.
PAAD domains are found in diverse proteins of unknown function and are structurally related to a superfamily of protein interaction modules that includes death domains, death effector domains, and Caspase activation and recruitment domains. Using bioinformatics strategies, cDNAs were identified that encode a novel protein of 110 kDa containing a PAAD domain followed by a putative nucleotide-binding (NACHT) domain and several leucine-rich repeat domains. This protein thus resembles Cryopyrin, a protein implicated in hereditary hyperinflammation syndromes, and was termed PAN2 for PAAD and NACHT-containing protein 2. When expressed in HEK293 cells, PAN2 suppressed NF-kappaB induction by the cytokines tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta), suggesting that this protein operates at a point of convergence in these two cytokine signaling pathways. This PAN2-mediated suppression of NF-kappaB was evident both in reporter gene assays that measured NF-kappaB transcriptional activity and electromobility shift assays that measured NF-kappaB DNA binding activity. PAN2 also suppressed NF-kappaB induction resulting from overexpression of several adapter proteins and protein kinases involved in the TNF or IL-1 receptor signal transduction, including TRAF2, TRAF6, RIP, IRAK2, and NF-kappaB-inducing kinase as well as the IkappaB kinases IKKalpha and IKKbeta. PAN2 also inhibited the cytokine-mediated activation of IKKalpha and IKKbeta as measured by in vitro kinase assays. Furthermore, PAN2 association with IKKalpha was demonstrated by co-immunoprecipitation assays, suggesting a direct effect on the IKK complex. These observations suggest a role for PAN2 in modulating NF-kappaB activity in cells, thus providing the insights into the potential functions of PAAD family proteins and their roles in controlling inflammatory responses.
Fiorentino
Loredana
L
Burnham Institute, La Jolla, California 92037, USA.
Stehlik
Christian
C
Oliveira
Vasco
V
Ariza
Maria Eugenia
ME
Godzik
Adam
A
Reed
John C
JC
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D024202:Electrophoretic Mobility Shift Assay; D004789:Enzyme Activation; D006801:Humans; D051550:I-kappa B Kinase; D007375:Interleukin-1 / Q000502:physiology*; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000096:biosynthesis*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D012097:Repressor Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology*
D048868:Adaptor Proteins, Signal Transducing; D017931:DNA Primers; D007375:Interleukin-1; D016328:NF-kappa B; C466775:NLRP4 protein, human; D011506:Proteins; D012097:Repressor Proteins; D014409:Tumor Necrosis Factor-alpha; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human