Fig. 3

TMPRSS2 isoform 1 downregulation induces SARS-CoV-2 replication arrest. a Schematic illustration of the SARS-CoV-2 non-infectious replicon [52] used in the following experiments. b Time course experiments of Caco-2 cells pre- treated 24 h with 1 μM CX-5461 or 10 μM PDS and electroporated with the SARS-CoV-2 non-infectious replicon [52]. Luciferase plate was read at time points 0, 12, 18, and 24 h. As a readout were obtained relative lights units (RLU) that reflect the replication of the replicon. c SARS-CoV-2 replication in Caco-2 cells transfected with siRNAs against TMPRSS2 isoform 1 and 2, only isoform 2 and scramble siRNA or treated 24 h with 10 and 25 μM camostat. Luciferase activity was measured 24 h after electroporation with the SARS-CoV2 non-infectious replicon [52]. d BG4-ChIP-qPCR experiment performed in Caco-2 cells untreated (light gray bars) or treated 24 h with 10 μM PDS (dark gray bars) or 1 μM CX-5461 (black bars). The values in the graph represent the ration between immunoprecipitated chromatin and DNA input. Significance was determined using an ordinary one-way ANOVA multiple comparison. Asterisks indicate statistical significance; in detail, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. e Caco-2 cells transfected with psiCHECK™-2 vector containing DNA oligonucleotides harboring the predicted G4 motifs at the exon 1 of the isoform 1, the G-rich sequence in the isoform 2 as well as control mutated sequences cloned upstream of the reporter gene. Expression of psiCHECK™-2 was normalized to the G4_exon1_Iso1 sample. f Luciferase assay in Caco-2 cells transfected with psiCHECK™-2 containing the predicted G4 motif at the exon 1 upstream of the reporter gene and either treated for 24 h with 1 μM CX-5461 or 10 μM PDS or left untreated. Expression of psiCHECK™-2 was normalized to the G4_exon1_Iso1 sample