Fig 1: Transduction with SARS-CoV-2 proteins increases apoptosis and causes cell cycle arrest in BEAS-2B cells.A) Representative immunoblots and densitometry showing upregulated expression of AIF and CDK4, 96 hours following transduction with 3 different SARS-CoV-2 proteins (vs GFP control). Only transduction with M protein downregulates CDK4. **P < 0.01; NS, not significant; n = 3/group.B) Representative images and mean data showing that heterologous expression of each of the three selected SARS-CoV-2 proteins increase the expression of AIF relative to the mitochondrial reporter protein, TOMM20, in BEAS-2B cells at 96 hours. Cells were stained with AIF mouse monoclonal antibody (green). Cells were also stained with TOMM20 rabbit polyclonal antibody (red). Ex/Em: 556/560-580 nm ***P < 0.001; n = 5–7/group.C) Lentiviral transduction of A549 cells with SARS-CoV-2 proteins, Nsp7, Nsp9 & M increase apoptosis, defined as cells that are annexin V positive and propidium iodide (PI) negative, relative to the GFP control. ***P < 0.001; n = 3/group.D) Lentiviral transduction of A549 cells with SARS-CoV-2 M protein causes cell cycle arrest in G1/G0, relative to GFP controls and other SARS-CoV-2 proteins. ***P < 0.001; NS; not significant; n = 3/group.E) BEAS-2B cells were loaded with mPTP staining dye-AM (Abcam ab239704; flow cytometry kit). CoCl2 was used to quench the cytosolic fluorescence, maintaining only the mitochondrial signal. Histograms demonstrate reduced mPTP activity (as the result of the mPTP opening) in M-transfected cells (light gray) versus control (dotted histogram). mPTP activity was not affected in Nsp9-transfected cells (dark gray) compared to control. The bar graph shows the fold change comparison in mPTP activity and confirms that that M-transfected cells had significantly reduced mPTP activity (open mPTP) versus control (p = 0.019; parametric one-way ANOVA). mPTP activity was calculated as MFI(CoCl2) – MFI (CoCl2+Ionomycin); MFI – median fluorescence intensity. MFI = mean fluorescent intensity. *P < 0.05; NS; not significant; n = 4/group.