PCIF1 facilitates viral entry into Calu-3 cells by regulating ACE2 and TMPRSS2 expression. A, Western blot analysis of ACE2 and TMPRSS2 protein levels in control or PCIF1-KO Calu-3 cells transfected with ACE2 and/or TMPRSS2 OE vectors. GAPDH served as a loading control. B, Luciferase assays quantifying infection of control or PCIF1-KO Calu-3 cells with luciferase-encoding SARS-CoV-2 (Left) or Omicron (Right) pseudoviruses. Data are presented as the mean ± SD (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001 by Student’s t test. C, Design of sgRNA-resistant (sgPCIF1-R) or inactive mutant (APPA) PCIF1 for rescue experiments. D, Western blot analysis of the indicated proteins in control or PCIF1-KO Calu-3 cells transfected with sgPCIF1-R or APPA OE vectors. E, Luciferase assay quantifying infection of the indicated Calu-3 cells with SARS-CoV-2 pseudovirus. Data are presented as the mean ± SD (n = 3). **P < 0.01 by Student’s t test.