Singleton-Merten syndrome (SMS) is an infrequently described autosomal-dominant disorder characterized by early and extreme aortic and valvular calcification, dental anomalies (early-onset ...periodontitis and root resorption), osteopenia, and acro-osteolysis. To determine the molecular etiology of this disease, we performed whole-exome sequencing and targeted Sanger sequencing. We identified a common missense mutation, c.2465G>A (p.Arg822Gln), in interferon induced with helicase C domain 1 (IFIH1, encoding melanoma differentiation-associated protein 5 MDA5) in four SMS subjects from two families and a simplex case. IFIH1 has been linked to a number of autoimmune disorders, including Aicardi-Goutières syndrome. Immunohistochemistry demonstrated the localization of MDA5 in all affected target tissues. In vitro functional analysis revealed that the IFIH1 c.2465G>A mutation enhanced MDA5 function in interferon beta induction. Interferon signature genes were upregulated in SMS individuals’ blood and dental cells. Our data identify a gain-of-function IFIH1 mutation as causing SMS and leading to early arterial calcification and dental inflammation and resorption.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPUK, ZAGLJ, ZRSKP
Mitochondria are descendants of endosymbiotic bacteria and retain essential prokaryotic features such as a compact circular genome. Consequently, in mammals, mitochondrial DNA is subjected to ...bidirectional transcription that generates overlapping transcripts, which are capable of forming long double-stranded RNA structures
. However, to our knowledge, mitochondrial double-stranded RNA has not been previously characterized in vivo. Here we describe the presence of a highly unstable native mitochondrial double-stranded RNA species at single-cell level and identify key roles for the degradosome components mitochondrial RNA helicase SUV3 and polynucleotide phosphorylase PNPase in restricting the levels of mitochondrial double-stranded RNA. Loss of either enzyme results in massive accumulation of mitochondrial double-stranded RNA that escapes into the cytoplasm in a PNPase-dependent manner. This process engages an MDA5-driven antiviral signalling pathway that triggers a type I interferon response. Consistent with these data, patients carrying hypomorphic mutations in the gene PNPT1, which encodes PNPase, display mitochondrial double-stranded RNA accumulation coupled with upregulation of interferon-stimulated genes and other markers of immune activation. The localization of PNPase to the mitochondrial inter-membrane space and matrix suggests that it has a dual role in preventing the formation and release of mitochondrial double-stranded RNA into the cytoplasm. This in turn prevents the activation of potent innate immune defence mechanisms that have evolved to protect vertebrates against microbial and viral attack.
Cellular innate immune sensors of DNA are essential for host defense against invading pathogens. However, the presence of self-DNA inside cells poses a risk of triggering unchecked immune responses. ...The mechanisms limiting induction of inflammation by self-DNA are poorly understood. BLM RecQ-like helicase is essential for genome integrity and is deficient in Bloom syndrome (BS), a rare genetic disease characterized by genome instability, accumulation of micronuclei, susceptibility to cancer, and immunodeficiency. Here, we show that BLM-deficient fibroblasts show constitutive up-regulation of inflammatory interferon-stimulated gene (ISG) expression, which is mediated by the cGAS-STING-IRF3 cytosolic DNA-sensing pathway. Increased DNA damage or down-regulation of the cytoplasmic exonuclease TREX1 enhances ISG expression in BLM-deficient fibroblasts. cGAS-containing cytoplasmic micronuclei are increased in BS cells. Finally, BS patients demonstrate elevated ISG expression in peripheral blood. These results reveal that BLM limits ISG induction, thus connecting DNA damage to cellular innate immune response, which may contribute to human pathogenesis.
Sensors of the innate immune system that detect intracellular nucleic acids must be regulated to prevent inappropriate activation by endogenous DNA and RNA. The exonuclease Trex1 regulates the ...DNA-sensing pathway by metabolizing potential DNA ligands that trigger it. However, an analogous mechanism for regulating the RIG-I-like receptors (RLRs) that detect RNA remains unknown. We found here that the SKIV2L RNA exosome potently limited the activation of RLRs. The unfolded protein response (UPR), which generated endogenous RLR ligands through the cleavage of cellular RNA by the endonuclease IRE-1, triggered the production of type I interferons in cells depleted of SKIV2L. Humans with deficiency in SKIV2L had a type I interferon signature in their peripheral blood. Our findings reveal a mechanism for the intracellular metabolism of immunostimulatory RNA, with implications for specific autoimmune disorders.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Considering that it is a single exon gene encoding a 314 amino acid protein, the genotype-phenotype landscape of
TREX1
is remarkably complex. Here we briefly describe the human diseases so-far ...associated with mutations in
TREX1
, which include Aicardi-Goutières syndrome, familial chilblain lupus, systemic lupus erythematosus and retinal vasculopathy with cerebral leukodystrophy.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
SAMHD1, an analogue of the murine interferon (IFN)-γ-induced gene Mg11 (ref. 1), has recently been identified as a human immunodeficiency virus-1 (HIV-1) restriction factor that blocks early-stage ...virus replication in dendritic and other myeloid cells and is the target of the lentiviral protein Vpx, which can relieve HIV-1 restriction. SAMHD1 is also associated with Aicardi-Goutières syndrome (AGS), an inflammatory encephalopathy characterized by chronic cerebrospinal fluid lymphocytosis and elevated levels of the antiviral cytokine IFN-α. The pathology associated with AGS resembles congenital viral infection, such as transplacentally acquired HIV. Here we show that human SAMHD1 is a potent dGTP-stimulated triphosphohydrolase that converts deoxynucleoside triphosphates to the constituent deoxynucleoside and inorganic triphosphate. The crystal structure of the catalytic core of SAMHD1 reveals that the protein is dimeric and indicates a molecular basis for dGTP stimulation of catalytic activity against dNTPs. We propose that SAMHD1, which is highly expressed in dendritic cells, restricts HIV-1 replication by hydrolysing the majority of cellular dNTPs, thus inhibiting reverse transcription and viral complementary DNA (cDNA) synthesis.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Objective
Type I interferon (IFN) is implicated in the pathogenesis of systemic lupus erythematosus (SLE) and interferonopathies such as Aicardi‐Goutières syndrome. A recently discovered ...DNA‐activated type I IFN pathway, cyclic GMP‐AMP synthase (cGAS), has been linked to Aicardi‐Goutières syndrome and mouse models of lupus. The aim of this study was to determine whether the cGAS pathway contributes to type I IFN production in patients with SLE.
Methods
SLE disease activity was measured by the Safety of Estrogens in Lupus Erythematosus National Assessment version of the Systemic Lupus Erythematosus Disease Activity Index. Expression of messenger RNA for cGAS and IFN‐stimulated genes (ISGs) was determined by quantitative polymerase chain reaction analysis. Cyclic GMP‐AMP (cGAMP) levels were examined by multiple reaction monitoring with ultra‐performance liquid chromatography tandem mass spectrometry.
Results
Expression of cGAS in peripheral blood mononuclear cells (PBMCs) was significantly higher in SLE patients than in normal controls (n = 51 and n = 20 respectively; P < 0.01). There was a positive correlation between cGAS expression and the IFN score (P < 0.001). The expression of cGAS in PBMCs showed a dose response to type I IFN stimulation in vitro, consistent with it being an ISG. Targeted measurement of cGAMP by tandem mass spectrometry detected cGAMP in 15% of the SLE patients (7 of 48) but none of the normal (0 of 19) or rheumatoid arthritis (0 of 22) controls. Disease activity was higher in SLE patients with cGAMP versus those without cGAMP.
Conclusion
Increased cGAS expression and cGAMP in a proportion of SLE patients indicates that the cGAS pathway should be considered as a contributor to type I IFN production. Whereas higher cGAS expression may be a consequence of exposure to type I IFN, detection of cGAMP in patients with increased disease activity indicates potential involvement of this pathway in disease expression.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
The patients, aged between 5 and 12 years, exhibited the phenotypic variability associated with TMEM173-activating mutations,2-4 with lung disease and systemic inflammation being the major features ...in patient 1 (P1) and patient 3 (P3), while in patient 2 (P2) skin involvement was most prominent (Fig 1; see Supplemental Text and Table E1 in this article's Online Repository at www.jacionline.org). Modest and incomplete downregulation of ISG was recently described in splenic B cells of mice treated with tofacitinib, a JAK1/3 inhibitor, with differential signaling effects suggesting currently poorly understood facets of IFN regulation.9 In this regard, our kinetic ex vivo experiments provide insights into the rapid dynamic changes in IFN signaling secondary to JAK1 blockade.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPUK, ZRSKP
Intracellular ISG15 is an interferon (IFN)-α/β-inducible ubiquitin-like modifier which can covalently bind other proteins in a process called ISGylation; it is an effector of IFN-α/β-dependent ...antiviral immunity in mice. We previously published a study describing humans with inherited ISG15 deficiency but without unusually severe viral diseases. We showed that these patients were prone to mycobacterial disease and that human ISG15 was non-redundant as an extracellular IFN-γ-inducing molecule. We show here that ISG15-deficient patients also display unanticipated cellular, immunological and clinical signs of enhanced IFN-α/β immunity, reminiscent of the Mendelian autoinflammatory interferonopathies Aicardi-Goutières syndrome and spondyloenchondrodysplasia. We further show that an absence of intracellular ISG15 in the patients' cells prevents the accumulation of USP18, a potent negative regulator of IFN-α/β signalling, resulting in the enhancement and amplification of IFN-α/β responses. Human ISG15, therefore, is not only redundant for antiviral immunity, but is a key negative regulator of IFN-α/β immunity. In humans, intracellular ISG15 is IFN-α/β-inducible not to serve as a substrate for ISGylation-dependent antiviral immunity, but to ensure USP18-dependent regulation of IFN-α/β and prevention of IFN-α/β-dependent autoinflammation.
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DOBA, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, SBMB, SIK, UILJ, UKNU, UL, UM, UPUK
In the RAS (renin-angiotensin system), Ang I (angiotensin I) is cleaved by ACE (angiotensin-converting enzyme) to form Ang II (angiotensin II), which has effects on blood pressure, fluid and ...electrolyte homoeostasis. We have examined the kinetics of angiotensin peptide cleavage by full-length human ACE, the separate N- and C-domains of ACE, the homologue of ACE, ACE2, and NEP (neprilysin). The activity of the enzyme preparations was determined by active-site titrations using competitive tight-binding inhibitors and fluorogenic substrates. Ang I was effectively cleaved by NEP to Ang (1-7) (kcat/K(m) of 6.2x10(5) M(-1) x s(-1)), but was a poor substrate for ACE2 (kcat/K(m) of 3.3x10(4) M(-1) x s(-1)). Ang (1-9) was a better substrate for NEP than ACE (kcat/K(m) of 3.7x10(5) M(-1) x s(-1) compared with kcat/K(m) of 6.8x10(4) M(-1) x s(-1)). Ang II was cleaved efficiently by ACE2 to Ang (1-7) (kcat/K(m) of 2.2x10(6) M(-1) x s(-1)) and was cleaved by NEP (kcat/K(m) of 2.2x10(5) M(-1) x s(-1)) to several degradation products. In contrast with a previous report, Ang (1-7), like Ang I and Ang (1-9), was cleaved with a similar efficiency by both the N- and C-domains of ACE (kcat/K(m) of 3.6x10(5) M(-1) x s(-1) compared with kcat/K(m) of 3.3x10(5) M(-1) x s(-1)). The two active sites of ACE exhibited negative co-operativity when either Ang I or Ang (1-7) was the substrate. In addition, a range of ACE inhibitors failed to inhibit ACE2. These kinetic data highlight that the flux of peptides through the RAS is complex, with the levels of ACE, ACE2 and NEP dictating whether vasoconstriction or vasodilation will predominate.
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