FIG 9.

Transcription factor Sp1 acted as the key regulator of HDAC1 transcription by binding to the HDAC1 promoter. (A) Expression of exogenous Sp proteins in IPEC-J2 cells transfected with pCMV-Sp1, -Sp2, -Sp3, or -Sp4 plasmids, shown by Western blotting using the anti-HA tag antibody. Vector control was included, and β-actin served as a loading control. (B) Transcription of HDAC1 by qRT-PCR in IPEC-J2 cells expressing different Sp proteins, as described for panel A. (C and D) HDAC1 expression in the IPEC-J2 cells treated with the Sp1/3 inhibitor mithramycin (MitA) by Western blotting (C) and qRT-PCR (D). Promoter binding of Sp1 (E) and Sp3 (F) in the cells transfected with pCMV-Sp1-HA or pCMV-Sp3-HA. The DNA fragments bound by Sp1 and Sp3 were pulled down by protein A/G beads preloaded with HA antibody for ChIP assay with mouse IgG as a control. (G) Sp1 knockdown efficiency of the uninfected and PEDV-infected cells transfected with Sp1 siRNA, detected by qRT-PCR. (H and I) PEDV N expression in the IPEC-J2 cells after Sp1 knockdown by qRT-PCR (H) and Western blotting (I). *, P < 0.05 for data in panels B, D, E, G, and H.