Fig 4.

BVDV downregulating mitochondrial GPX4 via inhibiting the protein levels of Nrf2. (A) Detection of intracellular GSH concentrations in mock-infected, CP BVDV (MOI = 5)-, and NCP BVDV (MOI = 10)-infected cells or erastin-treated cells with or without Fer-1 at 48 hpi. (B and C) Western blot analysis of Nrf2, SLC7A11, and GPX4 proteins in MDBK cells infected with CP BVDV (MOI = 5) (B) and NCP BVDV (MOI = 10) (C) at 24 and 48 hpi, respectively. (D and E) Western blot analysis of Nrf2, SLC7A11, and GPX4 proteins in MDBK cells infected with CP BVDV (MOI = 5) (D) and NCP BVDV (MOI = 10) (E) with or without Fer-1 at 48 hpi, respectively. (F) Establishment of MDBK cell lines stably overexpressing HA-Nrf2. Western blot analysis of GPX4 and SLC7A11 proteins in overexpressing HA-Nrf2 MDBK cells and NC MDBK cells infected with CP BVDV (MOI = 5) and NCP BVDV (MOI = 10) at 48 hpi, respectively. (G and H) Western blot analysis of GPX4 and VDAC1 proteins in cytoplasmic fractions (G) and mitochondrial fractions (H) from MDBK cells infected with CP BVDV (MOI = 5) at 24 and 48 hpi, respectively. (I and J) Western blot analysis of GPX4 and VDAC1 proteins in cytoplasmic fractions (I) and mitochondrial fractions (J) from MDBK cells infected with NCP BVDV (MOI = 10) at 24 and 48 hpi, respectively. (K and L) Western blot analysis of GPX4 and VDAC1 proteins in cytoplasmic fractions (K) and mitochondrial fractions (L) from MDBK cell lines stably overexpressing HA-Nrf2 infected with CP BVDV (MOI = 5) at 48 hpi, respectively. (M and N) Western blot analysis of GPX4 and VDAC1 proteins in cytoplasmic fractions (M) and mitochondrial fractions (N) from MDBK cell lines stably overexpressing HA-Nrf2 infected with NCP BVDV (MOI = 10) at 48 hpi, respectively. Data are given as means ± standard deviation from three independent experiments. P values were calculated using Student’s t test. An asterisk indicates a comparison with the indicated control. *, P < 0.05; **, P < 0.01; ns, not significant.