Journal: Molecular Neurodegeneration

Article Title: Mutations in PSEN1 predispose inflammation in an astrocyte model of familial Alzheimer’s disease through disrupted regulated intramembrane proteolysis

doi: 10.1186/s13024-025-00864-7

Figure Lengend Snippet: JAK-STAT signalling pathways are disrupted in PSEN1 mutant astrocytes. ( A-B ) FACs analysis of cell surface levels of IFNAR2 in CD44-positive iPSC-astrocytes, with and without TIC treatment. Data represent 5 experimental repeats from two batches and include 2 control lines and 2 PSEN1 mutant lines. Note that two control lines are pooled for TIC-treated datapoint. Data analysed within treatment subgroups via one-way ANOVA. ( C ) Representative Western blot of iPSC-astrocyte lysates with or without TIC treatment for 24 h. Actin is shown as a loading control. ( D-G ) Western blot analysis of total and phosphorylated MAPK (p42/p44), STAT2, STAT3 and NFκB under basal conditions or after 24 h TIC treatment. Note that one outlier was removed (grey arrow in panel A), due to a z score of 3.47. Data represent up to 4 independent batches and up to 7 technical repeats using 5 controls and 3 PSEN1 mutant lines (see Table ). For data separated by iPSC line, see Fig . ( H-J ) qPCR analysis of ISG15 , OAS1 and CXCL10 ; genes involved in interferon response (associated with JAK-STAT2 signalling). Note that CXCL10 was rarely detectable in untreated conditions (n.d.). Data represent up to 5 independent batches and up to 6 technical repeats from 6 control lines and 3 PSEN1 lines, see Table . For data separated by iPSC line, see Fig . ( K ) High content imaging analysis of nuclear NFκB normalised to cytosolic NFκB and plotted as a fold-change relative to maximum signal. Data represent 2 control lines and 3 PSEN1 lines, with two experimental repeats (see Table ). Individual data presented in Fig . Pairwise comparisons represent two tailed t-tests, where * = p < 0.05, ** = p < 0.01, *** = p < 0.001, and **** = p < 0.0001

Article Snippet: IFNAR2-APC , human , Miltenyi Biotec 130-099-560 , RRID: AB_2652223.

Techniques: Mutagenesis, Control, Western Blot, Imaging, Two Tailed Test

Journal: The Journal of Experimental Medicine

Article Title: A common form of dominant human IFNAR1 deficiency impairs IFN-α and -ω but not IFN-β-dependent immunity

doi: 10.1084/jem.20241413

Figure Lengend Snippet: Location of variants on human IFNAR1. (A) Schematic representation of full-length IFNAR1 protein, including the four fibronectin type III subdomains (SD1–4), the signal peptide (SP), and the transmembrane domain (TM). The mutations investigated in this study are depicted on the diagram. The hitherto unknown mutations are indicated in red, and the previously reported mutations are indicated in black. (B) Ribbon representation of the overall structure of the IFNα2–YNS–IFNAR1–IFNAR2 ternary complex (PDB 3SE3 , visualized with PyMOL [version 2.5.5]) showing IFNAR1 with SD1 colored in green, SD2 in blue, and SD3 in violet. IFNα2–YNS is depicted in cyan and IFNAR2 in beige. The amino-acid variants of IFNAR1 described here are highlighted by the depiction of their side chains as spheres across IFNAR1 SD1-3. Variants resulting from a missense mutation are depicted with side chains in yellow. Variants resulting in a complete LOF for IFN-α2, IFN-ω, and IFN-β signaling or hypomorphic for such signaling are depicted with side chains in magenta. In-frame indel variants are depicted with side chains in orange. Variants resulting from a frameshift mutation or an early stop codon (F45fs, W114X, T208fs, V225fs, and W261X) are not shown. (C) Close-up view of IFNAR1 SD1 (green) showing the location of variants N44 (orange), W73, and C79 (both magenta), along with V96 and the adjacent M128 from IFNAR1 SD2 (yellow). The locations of SD1 in relation to IFNAR1 SD2 (blue) and IFNα2-YNS (cyan) are shown. (D) Close-up view of IFNAR1 SD2 (blue), showing the location of P103, M128, and Y215 variants and the adjacent V96 from IFNAR1 SD1 (yellow), together with I144 (magenta). The locations of IFNAR1 SD2 (slate blue) in relation to IFNAR1 SD1 (green), IFNAR1 SD3 (violet), and IFNα2-YNS (cyan) are shown. (E) Close-up view of IFNAR1 SD3 (violet) showing the locations of the variants A264 (yellow) and S316 (magenta). The approximate locations of P334 (yellow circle) and P335 (orange circle), which were not resolved in the IFNα2–YNS–IFNAR1–IFNAR2 crystal structure, are predicted. The locations of IFNAR1 SD3 (violet) in relation to IFNAR1 SD2 (blue) and IFNα2-YNS (cyan) are shown.

Article Snippet: The cells were then stained by incubation with fluorescently labeled antibodies for 30–45 min at 4°C (mouse anti-IFNAR1: AA3 mAb [a gift from L. Runkel, Biogen, Inc.]; IFNAR2: Miltenyi Biotec, 130-128-948; HLA-I PE: R&D Systems, FAB7098P).

Techniques: Mutagenesis

Journal: The Journal of Experimental Medicine

Article Title: A common form of dominant human IFNAR1 deficiency impairs IFN-α and -ω but not IFN-β-dependent immunity

doi: 10.1084/jem.20241413

Figure Lengend Snippet: Expression of IFNAR1 by the patients’ fibroblasts. (A) IFNAR1 mRNA levels in SV40-fibroblasts from two healthy controls (C1, C2), and patients with IFNAR1 variants: P335del/P335del, P335del/+, W73C/W73C, V225fs/+, and V225fs/V225fs. GUS was used as an expression control. Graphs depict the mean ± SEM of two independent experiments, each with three technical duplicates. (B and C) Flow cytometry histograms of cell-surface expression for IFNAR1 (B) and IFNAR2 (C), with extracellular staining of SV40-fibroblasts from healthy controls and patients. Antibodies recognize the extracellular parts of IFNAR1 or IFNAR2. Each histogram plot is representative of two independent experiments.

Article Snippet: The cells were then stained by incubation with fluorescently labeled antibodies for 30–45 min at 4°C (mouse anti-IFNAR1: AA3 mAb [a gift from L. Runkel, Biogen, Inc.]; IFNAR2: Miltenyi Biotec, 130-128-948; HLA-I PE: R&D Systems, FAB7098P).

Techniques: Expressing, Control, Flow Cytometry, Staining