Transmissible Gastroenteritis Virus Infection Up-Regulates FcRn Expression via Nucleocapsid Protein and Secretion of TGF-ß in Porcine Intestinal Epithelial Cells

Qian, Shaoju; Gao, Zitong; Cao, Rui; Yang, Kang; Cui, Yijie; Li, Shaowen; Meng, Xianrong; He, Qigai; Li, Zili

Because NF-κB is a necessary transcription factor for FcRn production, we examined the effects of all TGEV-encoded proteins on NF-κB promoter activity. TGEV encodes 16 nonstructural proteins (nsp1-16), four structural proteins (S, E, M, and N), and three helper proteins (ORF7, ORF3a, and ORF3b), which were constructed recombinant plasmids by using the pCAGGS-HA vector. However, efficient expression occurred only for the plasmids encoding N, E, 3a, nsp1, nsp2, nsp5, nsp7, nsp8, nsp9, nsp10, nsp13, and nsp14 ( Figure 4C) . IPEC-J2 cells were co-transfected with NF-κB-Luc or FcRn, pRL-TK and different TGEV protein expression vectors. We found that TGEV N, nsp2 and nsp14 can all activate NF-κB (Figure 4A) , and TGEV N, 3a, FIGURE 2 | Involvement of TLR signaling cascades in FcRn activation by TGEV. (A) IPEC-J2 cells were transfected with 50 nM specific siRNAs targeting TLR2/3/4/8/9, TRIF, MyD88, RIG-I or NC siRNA, respectively for 24 h, then cells were collected for analysis of mRNA levels using RT-qPCR assays. (B) IPEC-J2 cells were transfected with the siRNAs TLR2/3/4/8/9, TRIF, MyD88, RIG-I or NC siRNA, respectively for 24 h, then with/without TGEV infection (MOI = 1). At 24 hpi, total RNA was extracted and FcRn or GAPDH mRNA expression was analyzed by RT-qPCR. (C,D) IPEC-J2 cells were transfected with the siRNAs TLR2/3/4/8/9, TRIF, MyD88, RIG-I or NC siRNA, respectively for 24 h, then with/without TGEV infection (MOI 1). IPEC-J2 cells were harvested at 36 hpi and measured by Western blot. The right panel represents the quantification of the bands by densitometry, corrected by the amount of GAPDH. * * p < 0.01. nsp1 and nsp5 can up-regulate FcRn by about 1.5-2-fold and N was the most significant inducer produced by FcRn (Figure 4B) .