Fig. 3: Effect of lead compounds on the expression of viral RNA and proteins. | Communications Biology

Fig. 3: Effect of lead compounds on the expression of viral RNA and proteins.

From: Targeting protein homeostasis with small molecules as a strategy for the development of pan-coronavirus antiviral therapies

Fig. 3

A Relative intracellular 229E RNA levels in 229E (MOI 1) infected Huh-7 cells after treatment with cordycepin, BTZ-1 and ADEP-42 at the indicated concentrations compared to DMSO for 24 hours. Genomic RNA (blue) was measured using primers against Orf1a, and total RNA (red) was measured using primers against N, both were normalized to actin mRNA. Mean ± SD shown from 2 independent experiments, each performed in duplicates. ****p < 0.0001. B Representative western blot of 229E N protein levels in samples treated as in A. Lysates were stained with anti-coronavirus N antibody and total protein was visualized using stain-free imaging. Molecular weight (MW) markers are given on the right in kDa. C Representative images of 229E S protein levels in samples treated as in A. Cells were fixed, processed for IF, and stained with anti-229E S antibody (green) and DAPI (blue). D Relative intracellular SARS-CoV-2 RNA levels in infected Huh-7 cells after treatment with cordycepin, BTZ-1 and ADEP-42 at 30 µM compared to DMSO for 48 hours. RNA was extracted and processed as in (A). Mean ± SD shown from 2 independent experiments, each performed in duplicate. ****p < 0.0001. E Representative western blot of SARS-CoV-2 N protein levels in samples treated as in D. The lysates were stained with anti-SARS-CoV-2 N antibody and GAPDH. MW markers are given on the right in kDa. F Representative images of SARS-CoV-2 S protein in infected Huh-7 cells after treatment with cordycepin at 30 µM, BTZ-1 at 10 µM, and ADEP-42 at 10 µM. Samples were fixed 48 hpi, processed for IF, and stained with anti-SARS-CoV-2 S antibody (green) and DAPI (blue). G Top: Effects of compounds on extracellular SARS-CoV-2 N RNA levels in infected (MOI 1) 3D lung organoids measured over days post infection (dpi). Bottom: Effects of compounds on cell viability as measured by LDH release over days post-treatment (dpt). Cordycepin (30 µM), BTZ-1 (10 µM) and ADEP-42 (10 µM) were compared with DMSO treatment. H Relative extracellular SARS-CoV-2 variants N RNA levels in media of infected Huh-7 cells after treatment with cordycepin (30 µM), BTZ-1 (10 µM) and ADEP-42 (10 µM) compared to DMSO.

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