Fig. 6: Progesterone and UPA are inhibitors of E-induced secretion and inflammation. | Nature Communications

Fig. 6: Progesterone and UPA are inhibitors of E-induced secretion and inflammation.

From: Coronavirus envelope protein activates TMED10-mediated unconventional secretion of inflammatory factors

Fig. 6

a Diagram of the high-throughput compound screening system based on cargo secretion analysis by complementary NanoLuc luciferase. b Heatmap showing the average mIL1β-HiBiT secretion levels from two independent experiments by each compound normalized to control. The numbers on the left side of heatmap show the serial numbers of compounds in each row. The heatmap shows the log2 value. c Quantification of relative mIL1β secretion levels in HEK293T cells in the presence of 10 μM indicated compounds. d–e mIL1β secretion in THP-1 cells expressed with Myc-tagged E of indicated coronaviruses in the presence of 10 μM UPA (d) or Progesterone (e) for 4 h. f Co-IP using HEK293T cells with TMED10-V5 and Myc-E-SARS2 in the absence or presence of 10 μM UPA or Progesterone. g Crosslink assay performed using HEK293T cells expressing TMED10-V5 without or with Myc-E-SARS2 in the absence or presence of 10 μM UPA or Progesterone. h Cell-free membrane translocation of mIL1β-FLAG in the absence or presence of Myc-E-SARS2, UPA or Progesterone (P4) as indicated. i–k C57BL/6 mice infected with AAV-Myc-E-SARS2 were intraperitoneal injected with or without UPA (1 mg/kg) followed by challenge with LPS and euthanized. Expression of Myc-E-SARS2 in the lung was verified by immunofluorescence (k, upper panel). Serum IL1β levels were determined by ELISA in (i). IL6 mRNA levels in indicated tissues were analyzed in (j). Lung inflammation was analyzed by H&E staining (k, lower panel) and inflammatory area was quantified in (l). m Chemical structure and names of Progesterone and its analogs with (Y) or without (N) inhibitory effect on mIL1β secretion in HEK293T cells shown in (n). n Quantification of relative mIL1β secretion levels in HEK293T cells in the absence or presence of compounds in (m). DSS, disuccinimidyl suberate; Prot K, Protease K; TX-100, TritonX-100. Data are mean ± s.d. Statistical significance was assessed one-way ANOVA (n = 4) followed by Dunnett’s multiple-comparison test (c, n), two-tailed t test (n = 5) (i, j, l). P values are indicated. Scale bars, 50 μm. Source data are provided as a Source Data file.

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