Fig. 2
From: UBXN3B positively regulates STING-mediated antiviral immune responses
UBXN3B is critical for STING-dependent IFN-I induction in mouse primary cells. a ELISA of IFN-α in the cell culture supernatants of (Ubxn3b+/+, Ubxn3b−/−) bone marrow-derived macrophages (Mϕ), Flt3-induced pDCs, and GM-CSF-induced cDCs (pooled from five littermates) 20 h after the indicated treatments. N = 3 per genotype. *P < 0.05 (unpaired Student’s t test). b Immunoblots of an interferon-stimulated gene Oas1a, Sting, and Ubxn3b expression in cDCs 20 h after the indicated treatments. Tubulin is a housekeeping protein control. qPCR analysis of (c) Ifnb1 and Tnfa mRNA expression and d cellular HSV-1 genome loads in cDCs infected with HSV-1 (MOI = 10) for the indicated time. e Immunoblots showing Ubxn3b and housekeeping Gapdh protein expression in mock (Ubxn3b+/+) or 4-hydroxyl tamoxifen-treated Cre+/− Ubxn3bflox/flox (Ubxn3b−/−) MEFs. f Viral titers (plaque-forming units/ml) in the supernatant of MEFs infected with HSV-1 (MOI = 0.1). N = 3 per genotype per time point. *P < 0.05; **P < 0.01 (unpaired Student’s t test). g qPCR analysis of selected immune gene mRNA expression in MEFs infected with HSV-1 as in g. h The immunoblots show knockout efficacy of STING and UBXN3B by CRISPR-Cas9 in human primary trophoblasts. Actin is a housekeeping protein control. i Fluorescent microscopic images of human primary trophoblasts infected with HSV-1-GFP (MOI = 0.3) for 18 h. Objective, ×5. Scale bar, 10 µm. j qPCR analysis of Ifnb1 mRNA expression in human primary trophoblasts infected with HSV-1-GFP for the indicated time. Bars/data points: mean ± s.e.m. Two biological replicates were pooled for qPCR (N = 2 per genotype per time point). *P < 0.05; **P < 0.01 (unpaired Student’s t test). The results are representative of two independent experiments
