Fig. 2.
m5C methyltransferase of NSUN2 is essential for its proviral activity. (A–E) Rescue of viral replication and gene expression by transfection of NSUN2 plasmid. NSUN2-KO or control A549 cells were transfected with 1 μg of empty vector (EV) pCAGGS or pCAGGS-NSUN2, followed by infection with rgRSV (A), hMPV (B), or VSV (C–E) at an MOI of 1.0. Transfection of pCAGGS-NSUN2 rescued RSV (A), hMPV (B), and VSV (E) protein expression, as well as GFP expression by rVSV-GFP (C) and VSV titer (D). (F–K) Two cysteine residues (C271 and C321) in NSUN2 protein are essential for its proviral activity. NSUN2-KO or control A549 cells were transfected with pCAGGS-NSUN2 or mutants followed by infection with rgRSV (F, G, and H) or rVSV-GFP (I, J, and K) at an MOI of 1.0. Transfection of WT NSUN2 but not NSUN2-C271A or NSUN2-C321A rescued the GFP expression (F and J), viral titer (G and K), and viral protein expression (H and I) in NSUN2-KO cells. All results are from three independent experiments. Data were analyzed using s Student’s t test (*P < 0.05; **P < 0.01).