Fig. 1: Spike protein of SARS-CoV-2 directly interacts with TLR4 and activates related immune responses.
From: SARS-CoV-2 spike protein interacts with and activates TLR41
a qRT-PCR analysis for the expression of IL1B in the THP-1 cells infected with 107 PFU/mL SARS-CoV-2 for 2 h, with or without 100 μM Resatorvid treatment. b Schematic representation of full-length spike protein and ectodomain (EC)-spike protein of SARS-CoV-2 (bottom), and affinity analysis of the binding of TLR4 to SARS-CoV-2 spike trimer (top). Purified SARS-CoV-2 spike trimer (SARS2-Spike-Tri, hereafter) was immobilized onto a CM5 sensor chip surface and tested for real-time association and dissociation of TLR4. c qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with control, 0.01 nM, 0.1 nM, 1 nM, 10 nM SARS2-Spike-Tri (top) or control, 0.5 ng/mL, 5 ng/mL, 50 ng/mL, 500 ng/mL LPS (bottom) for 12 h. d qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with control and Spike protein-pseudotyped (SPP) lentivirus for 12 h. e qRT-PCR analysis for the expression of IL1B in the control group and ATRA-differentiated HL-60 cells treated with 1 nM SARS2-Spike-Tri for 12 h. f qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with 10 nM Trimer Ectodomain (Tri), 10 nM N-terminal ___domain (NTD), 10 nM Receptor binding ___domain (RBD) of SARS-CoV-2 spike protein and control for 12 h. g qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with control, 1 nM SARS2-Spike-Tri, and 10 nM SARS2-Spike-Tri (left) or control, 50 ng/mL LPS, and 500 ng/mL LPS (right) for 2 h with or without 100 μM Resatorvid treatment. h qRT-PCR analysis for the expression of Il1b in WT and Myd88−/− Raw 264.7 cells treated with control, 500 ng/mL LPS, 10 nM SARS2-Spike-Tri for 6 h with or without 1 μM Resatorvid treatment. i qRT-PCR analysis for the expression of Il1b in WT and Tlr4−/− mouse peritoneal macrophage (top) or BMM (bottom) treated with control, 500 ng/mL LPS, 10 nM SARS2-Spike-Tri for 6 h. j qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with control, 10 nM SARS2-Spike-Tri, 10 nM SARS2-Spike-Tri + 0.025% Trypsin (1×), 10 nM SARS2-Spike-Tri + 0.25% Trypsin (10×) for 2 h. k qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with control, 1 × 105 PFU/mL, 1 × 106 PFU/mL, 1 × 107 PFU/mL, 1 × 108 PFU/mL MHV-A59 for 12 h. l qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with 1 × 107 PFU/mL MHV-A59 for 0 and 12 h with or without washing by PBS. m qRT-PCR analysis for the expression of IL1B in THP-1 cells treated with control, 10 nM SARS-Spike-Tri, 10 nM SARS-Spike-Tri + 100 μM Resatorvid for 2 h. SARS-Spike-Tri is short for the spike protein trimer of SARS-CoV. n qRT-PCR analysis for the expression of IL1B in THP-1 cells infected with 1 × 107 PFU/mL HCoV-229E for 2 h, with or without 100 μM Resatorvid treatment. o qRT-PCR analysis for the expression of Il1b in BMDMs of WT, Tlr2−/−, Tlr3−/− and Tlr4−/− mice. BMDMs were treated with control, 0.5 μg/mL Pam3CSK4, 20 μg/mL poly(I:C), 50 ng/mL LPS, 10 nM SARS2-Spike-Tri or 10 nM SARS-Spike-Tri for 2 h. p Volcano plots depicting the transcriptomes in THP-1 cells with or without 10 nM SARS2-Spike-Tri treatment for 2 h. All genes were shown with a false discovery rate (FDR) and fold change (FC) (red dots, FC > 1 and FDR < 0.05; green dots, FC < −1 and FDR < 0.05; black dots, −1 < FC < 1 or FDR > 0.05). q RNA-seq analysis for the upregulated interleukins (ILs) in THP-1 cells after 10 nM SARS2-Spike-Tri treatment for 2 h. The expression levels of each gene were showed by the fragments per kilobase of exons per million fragments mapped (FPKM). NS, non-significant; *P < 0.05; **P < 0.01; ***P < 0.001.
