Aberrant tRNA processing causes an autoinflammatory syndrome responsive to TNF inhibitors.

OBJECTIVES: To characterise the clinical features, immune manifestations and molecular mechanisms in a recently described autoinflammatory disease caused by mutations in TRNT1, a tRNA processing enzyme, and to explore the use of cytokine inhibitors in suppressing the inflammatory phenotype. METHODS: We studied nine patients with biallelic mutations in TRNT1 and the syndrome of congenital sideroblastic anaemia with immunodeficiency, fevers and developmental delay (SIFD). Genetic studies included whole exome sequencing (WES) and candidate gene screening. Patients' primary cells were used for deep RNA and tRNA sequencing, cytokine profiling, immunophenotyping, immunoblotting and electron microscopy (EM). RESULTS: We identified eight mutations in these nine patients, three of which have not been previously associated with SIFD. Three patients died in early childhood. Inflammatory cytokines, mainly interleukin (IL)-6, interferon gamma (IFN-γ) and IFN-induced cytokines were elevated in the serum, whereas tumour necrosis factor (TNF) and IL-1ß were present in tissue biopsies of patients with active inflammatory disease. Deep tRNA sequencing of patients' fibroblasts showed significant deficiency of mature cytosolic tRNAs. EM of bone marrow and skin biopsy samples revealed striking abnormalities across all cell types and a mix of necrotic and normal-appearing cells. By immunoprecipitation, we found evidence for dysregulation in protein clearance pathways. In 4/4 patients, treatment with a TNF inhibitor suppressed inflammation, reduced the need for blood transfusions and improved growth. CONCLUSIONS: Mutations of TRNT1 lead to a severe and often fatal syndrome, linking protein homeostasis and autoinflammation. Molecular diagnosis in early life will be crucial for initiating anti-TNF therapy, which might prevent some of the severe disease consequences.

Loss-of-function mutations in TNFAIP3 leading to A20 haploinsufficiency cause an early-onset autoinflammatory disease.

Systemic autoinflammatory diseases are driven by abnormal activation of innate immunity. Herein we describe a new disease caused by high-penetrance heterozygous germline mutations in TNFAIP3, which encodes the NF-κB regulatory protein A20, in six unrelated families with early-onset systemic inflammation. The disorder resembles Behçet's disease, which is typically considered a polygenic disorder with onset in early adulthood. A20 is a potent inhibitor of the NF-κB signaling pathway. Mutant, truncated A20 proteins are likely to act through haploinsufficiency because they do not exert a dominant-negative effect in overexpression experiments. Patient-derived cells show increased degradation of IκBα and nuclear translocation of the NF-κB p65 subunit together with increased expression of NF-κB-mediated proinflammatory cytokines. A20 restricts NF-κB signals via its deubiquitinase activity. In cells expressing mutant A20 protein, there is defective removal of Lys63-linked ubiquitin from TRAF6, NEMO and RIP1 after stimulation with tumor necrosis factor (TNF). NF-κB-dependent proinflammatory cytokines are potential therapeutic targets for the patients with this disease.

Recent advances in the molecular pathogenesis of hereditary recurrent fevers.

PURPOSE OF REVIEW: To discuss recent developments in the molecular basis of several hereditary recurrent fever syndromes, specifically the cryopyrin-associated periodic syndromes, familial Mediterranean fever and the tumor necrosis factor receptor associated periodic syndrome. RECENT FINDINGS: Mutations of CIAS1, the gene encoding cryopyrin/NALP3, lead to a spectrum of disease states termed the cryopyrinopathies. Recently, cryopyrin-deficient mice have been used to show that the protein is a key regulator of interleukin-1beta production that functions by recognizing stimuli such as bacterial RNA and infectious agents. Tumor necrosis factor receptor-associated periodic syndrome was initially thought to be caused by deficient metalloprotease-induced tumor necrosis factor receptor shedding, however new findings suggest that mutations in this receptor may result in inappropriate protein folding, leading to a host of other functional abnormalities that may cause inflammatory disease. Finally, data are emerging that address the possible function of the C-terminal B30.2 domain of pyrin, the familial Mediterranean fever protein. This motif has recently been shown to interact with and inhibit caspase-1, and the modeled structure of this complex highlights how mutations may affect the binding interface. SUMMARY: Recent reports have advanced our understanding of the structural and functional biology underlying the hereditary recurrent fevers, and are beginning to suggest possible mechanisms by which specific mutations cause disease.

Human transthyretin intronic open reading frames are not independently expressed in vivo or part of functional transcripts.

The human transthyretin (TTR) gene encodes a protein composed of four identical subunits with an important role in the plasma transport of thyroid hormone T4 and retinol. TTR spans 7.6 kilobases and consists of four exons. Two independent open reading frames (ORFs) with putative regulatory sequences have been described in the first and third introns, but their function--if any--is unknown. We have screened human cDNA libraries to determine if these sequences are transcribed. Transcripts of both ORFs were found in liver, pancreas and brain. Hybridization of the two sequences with multiple-tissue Northern blots further confirmed these results and revealed transcript sizes of approximately 1.5 and approximately 2.2 kb for ORF 1, and approximately 5.2 and approximately 7.8 kb for ORF 2. Rapid Amplification of cDNA Ends (RACE) was performed to characterize the full-length cDNAs containing each sequence. All products containing the ORFs were continuous in the genomic sequence corresponding to unspliced or partially spliced TTR. No evidence was found for novel transcripts containing productively spliced products of either ORF, or for shorter transcripts using the promoter and polyadenylation signals associated with them. ORF 1 RACE products identified in liver, pancreas and brain correspond to TTR transcripts in which intron 1 had not been removed; the transcripts containing ORF 2 may represent TTR hnRNA. Neither ORF is productively expressed as part of a larger transcript, or as an independent polypeptide.