An IKBKE variant conferring functional cGAS/STING pathway deficiency and susceptibility to recurrent HSV-2 meningitis
Azadeh Reyahi, Marie Studahl, Morten K. Skouboe, Stefanie Fruhwürth, Ryo Narita, Fanghui Ren, Moa Bjerhem Viklund, Marie B. Iversen, Mette Christiansen, Alexandra Svensson, Trine H. Mogensen, Kristina Eriksson, Søren R. Paludan
Azadeh Reyahi, Marie Studahl, Morten K. Skouboe, Stefanie Fruhwürth, Ryo Narita, Fanghui Ren, Moa Bjerhem Viklund, Marie B. Iversen, Mette Christiansen, Alexandra Svensson, Trine H. Mogensen, Kristina Eriksson, Søren R. Paludan
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Research Article Immunology Infectious disease

An IKBKE variant conferring functional cGAS/STING pathway deficiency and susceptibility to recurrent HSV-2 meningitis

Abstract

The mechanisms underlying susceptibility to recurrent herpes simplex virus type 2 (HSV-2) meningitis remain incompletely understood. In a patient experiencing multiple episodes of HSV-2 meningitis, we identified a monoallelic variant in the IKBKE gene, which encodes the IKKε kinase involved in induction of antiviral IFN genes. Patient cells displayed impaired induction of IFN-β1 (IFNB1) expression upon infection with HSV-2 or stimulation with double-stranded DNA (dsDNA) and failed to induce phosphorylation of STING, an activation marker of the DNA-sensing cyclic GMP-AMP synthase/stimulator of IFN genes (cGAS/STING) pathway. The patient allele encoded a truncated IKKε protein with loss of kinase activity and also capable of exerting dominant-negative activity. In stem cell–derived microglia, HSV-2–induced expression of IFNB1 was dependent on cGAS, TANK binding kinase 1 (TBK1), and IKBKE, but not TLR3, and supernatants from HSV-2–treated microglia exerted IKBKE-dependent type I IFN–mediated antiviral activity upon neurons. Reintroducing wild-type IKBKE into patient cells rescued IFNB1 induction following treatment with HSV-2 or dsDNA and restored antiviral activity. Collectively, we identify IKKε to be important for protection against HSV-2 meningitis and suggest a nonredundant role for the cGAS/STING pathway in human antiviral immunity.

Authors

Azadeh Reyahi, Marie Studahl, Morten K. Skouboe, Stefanie Fruhwürth, Ryo Narita, Fanghui Ren, Moa Bjerhem Viklund, Marie B. Iversen, Mette Christiansen, Alexandra Svensson, Trine H. Mogensen, Kristina Eriksson, Søren R. Paludan

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Figure 3

Impaired IFNB1 expression and increased viral load in HSV-2–infected P1 fibroblasts.

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Impaired IFNB1 expression and increased viral load in HSV-2–infected P1 ...
(A and B) Fibroblasts were infected with HSV-2. Total RNA was harvested after 6 and 24 hours and subjected to RT-qPCR for measurement of IFNA2 (A) and IFNB1 (B) mRNA. Cytokine mRNA levels were normalized to GAPDH, and the patient was compared with either 4 controls (for 6-hour stimulation) or 5 controls (for 24-hour infection). Unpaired t test was used for statistical analysis. (C) Fibroblasts were infected with HSV-1. Total RNA was harvested after 6 hours for IFNB1 mRNA quantification. (D) A plaque assay was used to measure HSV-2 titers in supernatants after 24 hours of infection. Nonparametric Mann-Whitney rank-sum test was used for statistical analysis. (E) Fibroblasts from P1 and 2 healthy controls were infected with measles virus (MeV) for 48 hours. A TCID50 (50% tissue culture infectious dose) assay was used to determine MeV titers in supernatants. Unpaired t test was used for statistical analysis. (F) Fibroblasts from P1 and patients functionally deficient in IRF3, IFNAR2, or TLR3 and fibroblasts from 2 healthy controls were infected with HSV-2 (MOI 1) for 24 hours, and a TCID50 assay was performed to quantify viral titers. Data were analyzed with 1-way ANOVA, and Dunnett’s multiple comparisons test was used for statistical analysis. Data shown in A and B are representative of 3 independent experiments, while the data in C represent 1 of 2 independent experiments. Data shown in D–F are pooled from 3 independent experiments. Error bars representing SEM and *, P ≤ 0.05; **, P ≤ 0.01; ****, P ≤ 0.0001.