Fig. 2: ARF1 acts as a pro-viral host factor for SARS-CoV-2 and its variants.

a–c HEK293 cells were co-transfected with the ACE2-Stag expression plasmid and ARF1-specific siRNAs or siRNA negative control (siNC). At 24 hpt, cells were infected with SARS-CoV-2 (MOI, 0.01) for 24 h. Following the confirmation of the ARF1 mRNA KD (a), SARS-CoV-2 propagation was then determined by plaque-forming unit (PFU) counting of infectious progeny virions (b). The viral N and ARF1 protein production was also analyzed by WB (c). d, e Wildtype (WT) or ARF1-KO (three clones) HEK293 cells were transfected with the ACE2 expression plasmid, followed by SARS-CoV-2 infection. At 24 hpi, the viral propagation (d) and protein production (e) were analyzed by plaque assays and WB, respectively. f–i The effects of ARF1 overexpression and reconstitution on SARS-CoV-2 propagation. WT cells (f, g) or the ARF1-KO (HEK293-ARF1-KO-2) cells (h, i) co-transfected with the ACE2 and ARF1 expression plasmids were similarly infected with SARS-CoV-2 at 24 hpt, followed by the viral propagation (f, h) and protein yield detections (g, i). j–m WT or ARF1-KO HEK293 cells transfected with the ACE2 expression plasmid were similarly infected with the indicated SARS-CoV-2 variants, followed by the viral propagation (j–l) and protein yield (m) analyses. Data are representative of three independent experiments with similar results (c, e, g, i, m). Graphs are presented as means ± SD, n = 3 (a, b, d, h, j–l) or 4 (f) independent biological replicates. Comparisons were performed with two-tailed Student’s t-test (f, h, j, k, l) or one-way ANOVA with Dunnett’s test (a, b, d). ^***p < 0.001; ^****p < 0.0001. See also Figs. S2 and S3. Source data with detailed p-values is provided as a Source data file.