Fig. 4

TLR3 can interact with NFY1 and CycTL in yeast two-hybrid and pull-down assays and can bind to the UGUAA, UGUAU and GGACU of ECT2 mRNA. a Yeast two-hybrid (Y2H) assay indicating that truncated TLR3 (∆TLR3, amino acids [aa] 1–111) interacts with NFY1 and CycTL. The bait and prey plasmids were co-transformed into yeast and positive colonies were selected on synthetic defined (SD) medium lacking Trp, Leu, His and Ade. b–d Pull-down assay indicating that AaTLR3 interacts with NFY1 (b), CycTL (c). Recombinant GST-TLR3, His-NFY1 and His-CycTL were produced in E. coli Rosetta (DE3) and purified. After incubation in vitro, the protein mixture was separated by SDS-PAGE gel. Proteins were detected with anti-His and anti-GST antibodies. d, e ECT2-His interacts with the UGUAA, UGUAU and GGACU of TLR3 mRNA on RNA-EMSA. Recombinant ECT2-His was produced in E. coli Rosetta (DE3) and purified. The concentration ratios of free probe and bound probe in the third to sixth lane were 1:1, 1:10, 1:100 and 1:1000, respectively