The inflammasome.
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Adaptor Proteins, Signal Transducing↗
Biomedical subjects
Stéphanie Papin
Publications and source records attributed to Stéphanie Papin.
Interaction of pyrin with 14.3.3 in an isoform-specific and phosphorylation-dependent manner regulates its translocation to the nucleus.
OBJECTIVE: Pyrin, the familial Mediterranean fever gene product, exists in several isoforms of unknown functions. The recombinant full-length isoform (pyrin.fl) is cytoplasmic, whereas an alternatively spliced isoform lacking exon 2 (pyrin.DeltaEx2) concentrates in the nucleus. Native pyrin, mainly consisting of pyrin.fl, is also cytoplasmic in monocytes but is predominantly nuclear in other cell types. To understand pyrin-dependent biologic pathways and to decipher the mechanisms accounting for such different patterns of subcellular compartmentalization, binding partners and posttranslational modifications of pyrin were assessed. METHODS: A yeast 2-hybrid screen was performed with pyrin.fl as the bait. The interaction identified between pyrin.fl and 14.3.3 proteins was confirmed by immunoprecipitation of (35)S-radiolabeled protein complexes; similar experiments were performed with pyrin.DeltaEx2, pyrin.fl after alkaline phosphatase treatment, and pyrin.fl mutants in which several exon 2-encoded serine residues were replaced by nonphosphorylatable alanines. The subcellular localization of the different wild-type and mutated pyrin proteins was assessed by immunofluorescence. RESULTS: Two members of the 14.3.3 protein family were identified as pyrin partners. Whereas pyrin.fl interacted with 14.3.3tau and 14.3.3epsilon, these interactions did not occur with pyrin.DeltaEx2. Pyrin.fl was phosphorylated, and this modification mediated 14.3.3 binding. Serines 208, 209, and 242, within exon 2, acted as critical residues in the interaction between pyrin.fl and 14.3.3. When an S208-S209-S242A pyrin.fl triple mutant or wild-type pyrin.fl in the presence of an inhibitor of 14.3.3-ligand interactions was used, promotion of nuclear translocation of pyrin was observed. CONCLUSION: These results disclose the role played by 14.3.3 in the regulation of the subcellular compartmentalization of pyrin in a phosphorylation- and isoform-dependent manner. They also reconcile the observations made in vitro with those made in vivo, while providing a direct link between 14.3.3-dependent pathways and pyrin.
Cell Nucleus↗
The tumor necrosis factor alpha-dependent activation of the human mediterranean fever (MEFV) promoter is mediated by a synergistic interaction between C/EBP beta and NF kappaB p65.
MEFV is a gene expressed specifically in myeloid cells and whose mutations underlie an autosomal recessive auto-inflammatory disease, called familial Mediterranean fever (FMF), characterized by recurrent episodes of serosal inflammation. This gene, which encodes a protein with unclear physiological functions, has been shown to be up-regulated by the pro-inflammatory cytokine tumor necrosis factor alpha (TNFalpha). However, the mechanism of this regulation is unknown, and the MEFV promoter is still to be characterized. Here, we show that 243 bp of the 5'-flanking region of the human MEFV gene are sufficient to direct high level expression of MEFV in TNFalpha-treated cells. The TNFalpha-induced expression of MEFV is dependent on both NFkappaB p65 and C/EBPbeta that bind to evolutionarily conserved sites located, in the human promoter, at positions -163 and -55, respectively. As shown by a series of transcription and gel shift assays performed with wild-type and mutated promoter sequences, these two transcription factors act differently on the TNFalpha-dependent transcription of MEFV: C/EBPbeta is the key regulatory factor required to confer cell responsiveness to TNFalpha, whereas NFkappaB p65 increases this response by means of a synergistic interaction with C/EBPbeta that is dependent on the integrity of the identified -55 C/EBP binding site. Given the phenotype of patients with FMF, this C/EBP-NFkappaB interaction may represent a key step in the control of an inflammatory response that is abnormally high in this disease. These data, which shed novel light on the pathophysiology of FMF, represent an unusual example of cross-talk between C/EBP and NFkappaB pathways in TNFalpha signaling.
Base Sequence↗
Familial Mediterranean fever among patients from Karabakh and the diagnostic value of MEFV gene analysis in all classically affected populations.
OBJECTIVE: Familial Mediterranean fever (FMF) is an autosomal-recessive disorder that is common in Armenian, Turkish, Arab, and Sephardic Jewish populations. Its clinical diagnosis is one of exclusion, with the patients displaying nonspecific symptoms related to serosal inflammation. MEFV gene analysis has provided the first objective diagnostic criterion for FMF. However, in the absence of an identified mutation (NI/NI genotype), both the sensitivity of the molecular analyses and the involvement of the MEFV gene in FMF are called into question. The present study was designed to further evaluate the diagnostic value of MEFV analysis in another population of Mediterranean extraction. METHODS: The MEFV gene was screened for mutations in 50 patients living in Karabakh (near Armenia) who fulfilled the established criteria for FMF. In addition, we analyzed published series of patients from the above-mentioned at-risk populations. RESULTS: The mutation spectrum in Karabakhian patients, which consisted of only 6 mutations (with 26% of NI alleles), differed from that reported in Armenian patients. Strikingly, among patients from Karabakh and among all classically affected populations, the distribution of genotypes differed dramatically from Hardy-Weinberg equilibrium (P = 0.0016 and P < 0.00001, respectively). These results, combined with other population genetics-based data, revealed the existence of an FMF-like condition that, depending on the patients' ancestry, was shown to affect 85-99% of those with the NI/NI genotype. CONCLUSION: These data illuminate the meaning of negative results of MEFV analyses and show that in all populations evaluated, most patients with the NI/NI genotype had disease that mimicked FMF and was unrelated to the MEFV gene. Our findings also demonstrate the high sensitivity of a search for very few mutations in order to perform a molecular diagnosis of MEFV-related FMF.
Adolescent↗