Figure 1

1. Cellular effects in host cells induced by recombinant VP1 (rVP1) of foot-and-mouth disease virus (FMDV) Chung-Da Yang, Jei-Ming Peng, Shu-Mei Liang Institute of BioAgricultural Sciences, Academia Sinica, Taipei, Taiwan Figure 6 Figure 3 Introduction - 2h 4h 6h 8h 1h 0.5 M rVP1 (a) Foot-and-mouth disease virus (FMDV) is an etiological agent of foot-and-mouth disease (FMD). Capsid protein VP1 of FMDV contains a highly conserved Arg-Gly-Asp (RGD) tripeptide, which binds to integrins and facilitates the viral internalization into target cells. We cloned and purified the recombinant VP1 (rVP1) in an aqueous form previously. In this study, we investigated the cellular effects induced by rVP1 in BHK-21 and RAW 264.7 cells. 0.1 M rVP1 0.5 M rVP1 2.0 M rVP1 - TLR1 1h 8h 4h 8h 1h 4h 8h 4h 1h TLR2 TLR9 TLR3 -actin TLR4 TLR5 (b) - - TLR6 + + Anti-51 integrin Ab - - TLR7 Figure 1 + + 0.5 M rVP1 TLR8 TLR9 TLR9 -actin -actin rVP1 rVP1 treatment induced temporal reduction of TLR9 (a) and the reduction was reversed by anti-51 integrin antibody. Cell Cellular effects? rVP1 treatment is specific to reduce TLR9 expression. Figure 7 - 1h 2h 4h 8h 30’ 0.5 M rVP1 Figure 4 TLR9 Phospho-p38 Phospho-ERK1/2 Total p38 Effects of rVP1 on MAPKs phosphorylation. 1h 2h 8h 4h rVP1 treatment caused apoptosis and this effect was reversed by anti-VP1 and anti-α5β1 antibodies as well as fibronectin. Summary pNF-B  + + + + + • Recombinant VP1 (rVP1) of FMDV induced apoptosis in BHK-21 cells after binding to integrins. • rVP1 reduced the expression of toll-like receptor 9 (TLR9), an immune sensor that recognized bacterial and virus DNA. • The reduction of TLR9 expression by rVP1 was at both translational and transcriptional levels. • The reduction of TLR9 expression by rVP1 could be reversed by PDTC, NF-B inhibitor, suggesting that NF-B involved in expression of TLR9 regulated by rVP1. • Effects of rVP1 on MAPKs phosphorylation implied that ERK1/2 and p38 MAPKs might be involved in reduction of TLR9 by rVP1. • These results suggested that rVP1 may reduce host defensive response to viral invasion and trigger apoptosis in host cells. 0.5 M rVP1   + + + + Figure 2 rVP1 treatment induced NF-B activity. (a) BHK-21 cells - 1h 0.5 M rVP1 2h 4h 6h 8h TLR9 Figure 5 -actin - - + + 30 M PDTC (b) - RAW 264.7 cells - + - + 0.5 M rVP1 0.5 M rVP1 1h 2h 4h 6h 8h TLR9 TLR9 -actin -actin PDTC, NF-B inhibitor, reversed the reduction of TLR9 by rVP1. rVP1 treatment caused TLR9 reduction in BHK-21 (a) and RAW 264.7 (b) cells.