Extended Data Fig. 5: SARS-CoV-2 induces NEMO cleavage.
a, SARS-CoV-2- (MOI: 1) or mock-infected Vero E6 cells were harvested at the indicated time points. Cell lysates were immunoblotted for NEMO. N = 3 experiments. b, Human brain endothelial hCMEC/D3 cells were transfected with plasmids encoding for NEMO-2A and for human ACE2 plus TMPRSS2. Twenty-four hours after transfection, cells were incubated with SARS-CoV-2 for 2 hours (MOI: 1) and harvested after additional 24 hours. Only transfected cells expressing human ACE2 as well as 2A-tagged NEMO were susceptible to SARS-CoV-2 infection (Fig. 2e). Accordingly, 2A-tagged NEMO but not untagged NEMO was degraded. Upper panel, immunoblot using anti-NEMO; lower panel, immunoblot using anti-2A antibodies. N = 3 experiments. c, Lysates of the medulla oblongata of SARS-CoV-2-infected and control patients were immunoblotted for NEMO. Samples were matched according to post-mortem intervals (PMI): lanes 1 and 2: 1 day PMI; lanes 3 and 4: 2–3 days PMI; lanes 5 and 6: 4–5 days PMI; lanes 7 and 8: 4–5 days PMI. Histogram intensities of each lane were measured and depicted as means ± s.e.m., showing a decrease of the native forms of NEMO and an increase in cleaved NEMO in the tissue of SARS-CoV-2-infected patients (red profile) in comparison to control subjects (black profile). N = 4 patients in each group.
