Background/Purpose: Mutations in the TNFRSF1A, which encodes tumor necrosis factor receptor 1 (TNFR1), are associated with the autosomal dominant disease TNF Receptor- Associated Periodic Syndrome (TRAPS). TRAPS is an autoinflammatory disease characterized by intermittent self-limited inflammatory episodes of fever. More than 100 heterozygous TNFRSF1A mutations associated with TRAPS or TRAPS-like clinical phenotype have been reported. T50M mutation and cysteine mutations are reported to cause intracellular accumulation of TNFR1 and exhibit a severe disease phenotype. Single nucleotide polymorphism in TNFR1, such as T61I and R92Q mutations, occurs in 1-5% of the general population and can be associated with a TRAPS-like phenotype. Therefore, there are multiple responsible mechanisms to induce inflammation in patients with TRAPS, and the molecular pathogensis is not yet fully understood. Recently we have identified a novel mutation, G58V (p.G87V) in TNFRSF1A, in two individuals in a family. They also had T61I mutation, and had suffered from recurrent episodes of intermittent fever and abdominal pain. In this study, we functionally characterized this novel G58V TNFRSF1A mutation.

Methods: The possible pathogenicity of the G58V mutation was analyzed using the four online prediction tools (SIFT, Polyphen2, PROVEAN and PANTHER). Wild-type (WT) or mutated TNFRSF1A (T50M, G58V, T61I, R92Q) constructs were generated by cloning the individual cDNAs into the pcDNA3.1 vector. The TNFR1 constructs were transfected into HEK-293 cells. Expression levels of the TNFR1 were examined by western blotting. To evaluate the NF-κB promoter activity, HEK-293 cells were transfected with the TNFR1 constructs along with NF-κB promoter-driven luciferase plasmid and the secreted alkaline phosphatase control vector pSEAP. NF-κB activation levels at 24 hours after the transfection were measured by the dual-luciferase reporter assay.

Results: The novel G58V mutation was predicted to be a highly damaging amino acid substitution that could cause a disease. The result is similar to those of pathogenic T50M and cysteine mutations. Expression levels of the WT and mutated TNFR1 proteins are comparable in the whole cell lysates of HEK-293 cells. NF-κB promoter activities in the T50M or the G58V TNFR1-expressing cells were significantly decreased compared to those in WT TNFR1-expressing cells (mean±SD; T50M 29.6±7.5% vs. WT 100±11.5%, p<0.001; G58V 40.1±0.9% vs. WT, p<0.001). The T61I mutation showed a small but significant reducation in NF-κB promoter activity (67.4±18.8%, p<0.05 vs. WT). In contrast, the R92Q mutation did not suppress NF-κB promoter activity (93.2±18.4%).

Conclusion: Consistent with a previous report (Blood 2006;108:1320-1327), the T50M mutation suppressed spontaneous NF-κB promoter activity, in spite of the clinical inflammatory features of TRAPS. The newly identified G58V mutation developed the similar phenotype to the pathogenic T50M mutation. These findings indicate that G58V could be a responsible mutation causing TRAPS.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/functional-analysis-of-the-novel-g58v-mutation-in-the-tnfrsf1a-gene-identified-in-a-family-with-tnf-receptor-associated-periodic-syndrome-traps/