Victoria Tüngler

Defective removal of ribonucleotides from DNA promotes systemic autoimmunity

Genome integrity is continuously challenged by the DNA damage that arises during normal cell metabolism. Biallelic mutations in the genes encoding the genome surveillance enzyme ribonuclease H2 (RNase H2) cause Aicardi-Goutières syndrome (AGS), a pediatric disorder that shares features with the autoimmune disease systemic lupus erythematosus (SLE). Here we determined that heterozygous parents of AGS patients exhibit an intermediate autoimmune phenotype and demonstrated a genetic association between rare RNASEH2 sequence variants and SLE. Evaluation of patient cells revealed that SLE- and AGS-associated mutations impair RNase H2 function and result in accumulation of ribonucleotides in genomic DNA. The ensuing chronic low level of DNA damage triggered a DNA damage response characterized by constitutive p53 phosphorylation and senescence. Patient fibroblasts exhibited constitutive upregulation of IFN-stimulated genes and an enhanced type I IFN response to the immunostimulatory nucleic acid polyinosinic:polycytidylic acid and UV light irradiation, linking RNase H2 deficiency to potentiation of innate immune signaling. Moreover, UV-induced cyclobutane pyrimidine dimer formation was markedly enhanced in ribonucleotide-containing DNA, providing a mechanism for photosensitivity in RNase H2-associated SLE. Collectively, our findings implicate RNase H2 in the pathogenesis of SLE and suggest a role of DNA damage-associated pathways in the initiation of autoimmunity.

Single-stranded nucleic acids promote SAMHD1 complex formation

SAM domain and HD domain-containing protein 1 (SAMHD1) is a dGTP-dependent triphosphohydrolase that degrades deoxyribonucleoside triphosphates (dNTPs) thereby limiting the intracellular dNTP pool. Mutations in SAMHD1 cause Aicardi–Goutières syndrome (AGS), an inflammatory encephalopathy that mimics congenital viral infection and that phenotypically overlaps with the autoimmune disease systemic lupus erythematosus. Both disorders are characterized by activation of the antiviral cytokine interferon-α initiated by immune recognition of self nucleic acids. Here we provide first direct evidence that SAMHD1 associates with endogenous nucleic acids in situ. Using fluorescence cross-correlation spectroscopy, we demonstrate that SAMHD1 specifically interacts with ssRNA and ssDNA and establish that nucleic acid-binding and formation of SAMHD1 complexes are mutually dependent. Interaction with nucleic acids and complex formation do not require the SAM domain, but are dependent on the HD domain and the C-terminal region of SAMHD1. We finally demonstrate that mutations associated with AGS exhibit both impaired nucleic acid-binding and complex formation implicating that interaction with nucleic acids is an integral aspect of SAMHD1 function.

Familial chilblain lupus due to a gain-of-function mutation in STING

Objectives Familial chilblain lupus is a monogenic form of cutaneous lupus erythematosus caused by loss-of-function mutations in the nucleases TREX1 or SAMHD1. In a family without TREX1 or SAMHD1 mutation, we sought to determine the causative gene and the underlying disease pathology. Methods Exome sequencing was used for disease gene identification. Structural analysis was performed by homology modelling and docking simulations. Type I interferon (IFN) activation was assessed in cells transfected with STING cDNA using an IFN-β reporter and Western blotting. IFN signatures in patient blood in response to tofacitinib treatment were measured by RT-PCR of IFN-stimulated genes. Results In a multigenerational family with five members affected with chilblain lupus, we identified a heterozygous mutation of STING, a signalling molecule in the cytosolic DNA sensing pathway. Structural and functional analyses indicate that mutant STING enhances homodimerisation in the absence of its ligand cGAMP resulting in constitutive type I IFN activation. Treatment of two affected family members with the Janus kinase (JAK) inhibitor tofacitinib led to a marked suppression of the IFN signature. Conclusions A heterozygous gain-of-function mutation in STING can cause familial chilblain lupus. These findings expand the genetic spectrum of type I IFN-dependent disorders and suggest that JAK inhibition may be of therapeutic value.

Inherited or de novo mutation affecting aspartate 18 of TREX1 results in either familial chilblain lupus or Aicardi-Goutières syndrome.

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Naturally Occurring Genetic Variants of Human Caspase‐1 Differ Considerably in Structure and the Ability to Activate Interleukin‐1β

Caspase‐1 (Interleukin‐1 Converting Enzyme, ICE) is a proinflammatory enzyme that plays pivotal roles in innate immunity and many inflammatory conditions such as periodic fever syndromes and gout. Inflammation is often mediated by enzymatic activation of interleukin (IL)‐1β and IL‐18. We detected seven naturally occurring human CASP1 variants with different effects on protein structure, expression, and enzymatic activity. Most mutations destabilized the caspase‐1 dimer interface as revealed by crystal structure analysis and homology modeling followed by molecular dynamics simulations. All variants demonstrated decreased or absent enzymatic and IL‐1β releasing activity in vitro, in a cell transfection model, and as low as 25% of normal ex vivo in a whole blood assay of samples taken from subjects with variant CASP1, a subset of whom suffered from unclassified autoinflammation. We conclude that decreased enzymatic activity of caspase‐1 is compatible with normal life and does not prevent moderate and severe autoinflammation.

Effect of intestinal pressure on fistula closure during vacuum assisted treatment: A computational approach

BACKGROUND Enterocutaneous fistulae, pathological communications between the intestinal lumen and the abdominal skin, can arise as serious complication of gastrointestinal surgery. A current non-surgical treatment for this pathology involves topical application of sub-atmospheric pressure, also known as vacuum assisted closure (VAC). While this technique appears to be promising, surgeons report a number of cases in which its application fails to achieve fistula closure. Here, we evaluate the fistulas physical properties during the vacuum assisted closure process in a computational approach exploring the relevance of intraluminal intestinal pressure. METHODS A mathematical model formulated by differential equations based on tissue elasticity properties and principles of fluid mechanics was created and forcing functions were integrated to mimic intestinal pressure dynamics. A software to solve equations and to fit the model to experimentally obtained data was developed. This enabled simulations of vacuum assisted fistula closure under different intestinal pressure. RESULTS The simulation output indicates conditions, in which fistula closure can or cannot be expected suggesting favoured or impeded healing, respectively. When modifications of intestinal pressure, as observed in fistula accompanying pathologies, are integrated, the outcome of fistula closure changes considerably. Rise of intestinal pressure is associated with delay of fistula closure and temporary fistula radius augmentation, while reduction of intestinal pressure during sub-atmospheric pressure treatment contributes to a faster and direct fistula closure. CONCLUSION From the model predictions, we conclude that administration of intestinal pressure decreasing compounds (e.g. butylscopolamine, glucagon) may improve VAC treatment, while intestinal pressure increasing drugs should be avoided.

Response to: ‘JAK inhibition in STING-associated interferonopathy’ by Crow et al

We appreciate the comments by Crow et al on our report of a gain-of-function mutation in STING in familial chilblain lupus and the effect of the JAK inhibitor tofacitinib.1 In a concurrent study,2 Crow et al reported a marked clinical improvement in three patients with stimulator of interferon genes (STING)-associated vasculopathy3 using the JAK inhibitor ruxolitinib. This was accompanied by an incomplete reduction in the expression of interferon (IFN)-stimulated genes (ISGs) in blood raising the question as to the IFN signature as a read-out of clinical efficacy. This is a very important question that we cannot answer with regard to tofacitinib as we have no long-term experience with this JAK inhibitor in patients with type I interferonopathies. However, we have used ruxolitinib in two patients with Aicardi-Goutieres syndrome (AGS), an inflammatory encephalopathy characterised by chronic type I IFN activation.4 Both patients carried biallelic RNASEH2B mutations and suffered from severe developmental delay. They received no other medications, when treatment with ruxolitinib …

Variable clinical phenotype in two siblings with Aicardi-Goutières syndrome type 6 and a novel mutation in the ADAR gene

Aicardi-Goutières syndrome (AGS) is a hereditary inflammatory encephalopathy resulting in severe neurological damage in the majority of cases. We report on two siblings with AGS6 due to compound heterozygosity for a known and a novel mutation in the ADAR gene and a strikingly variable phenotype. The first sibling presented at 12 months of age with a subacute encephalopathy following a mild respiratory infection. The child developed a spastic tetraparesis, generalized dystonia and dysarthria. In contrast, the younger sibling presented with an acute episode of neurological impairment in his third year of life, from which he recovered without sequelae within a few weeks. These findings illustrate a striking intrafamilial phenotypic variability in patients with AGS6 and describe the first case of a full recovery from an acute encephalopathy in an AGS patient. Our findings also suggest that AGS should be considered as an important differential diagnosis of an infection-triggered encephalopathy in infancy despite the absence of typical neuroimaging findings.

Therapeutic Approaches to Type I Interferonopathies

Purpose of ReviewTo review recent scientific advances and therapeutic approaches in the expanding field of type I interferonopathies.SummaryType I interferonopathies represent a genetically and phenotypically heterogenous group of disorders of the innate immune system caused by constitutive activation of antiviral type I interferon (IFN). Clinically, type I interferonopathies are characterized by autoinflammation and varying degrees of autoimmunity or immunodeficiency. The elucidation of the underlying genetic causes has revealed novel cell-intrinsic mechanisms that protect the organism against inappropriate immune recognition of self nucleic acids by cytosolic nucleic acid sensors. The type I IFN system is subject to a tight and complex regulation. Disturbances of its checks and balances can spark an unwanted immune response causing uncontrolled type I IFN signaling. Novel mechanistic insight into pathways that control the type I IFN system is providing opportunities for targeted therapeutic approaches by repurposing drugs such as Janus kinase inhibitors or reverse transcriptase inhibitors.

Klinische Symptome und Pathogenese der Typ-1-Interferonopathien

ZusammenfassungDie Typ-1-Interferonopathien stellen eine Gruppe genetisch bedingter seltener Erkrankungen dar, die durch eine Fehlfunktion des angeborenen Immunsystems hervorgerufen werden. Gemeinsames Kennzeichen ist eine Dysregulation der antiviralen Typ-1-Interferon(IFN)-Achse, die zu einer konstitutiven Typ-1-IFN-Aktivierung führt. Als systemisch-entzündliche Erkrankungen ist das klinische Spektrum der Typ-1-Interferonopathien sehr breit und sowohl durch Autoinflammation als auch durch Autoimmunität gekennzeichnet. Hierbei stehen neurologische und kutane Manifestationen im Vordergrund. Pathogenetisch liegen den Typ-1-Interferonopathien Störungen im Metabolismus und in der immunologischen Erkennung von intrazellulären Nukleinsäuren zugrunde. Unser derzeitiges Verständnis der molekularen Pathogenese der Typ-1-Interferonopathien weist darauf hin, dass eine immunmodulatorische Intervention, die der inadäquaten Typ-1-IFN-Aktvierung entgegenwirkt, therapeutisch wirksam sein könnte.AbstractType 1 interferonopathies represent a group of genetically determined rare diseases caused by defects of the innate immune system. Central to all type 1 interferonopathies is a dysregulation of the antiviral type 1 interferon (IFN) axis, which results in constitutive overproduction of type 1 IFN. All type 1 interferonopathies present as systemic inflammatory disorders characterized by autoinflammation and autoimmunity. Although the clinical spectrum is highly variable and broad, neurological and cutaneous manifestations represent the most salient findings. Chronic type 1 IFN activation is due to defects in pathways affecting the metabolism or the immune recognition of intracellular nucleic acids. The current understanding of the molecular mechanisms underlying type 1 interferonopathies indicates that an immunomodulatory intervention targeting the type 1 IFN axis might be of therapeutic value.