Effect of non-specific RNA-binding peptide on BiFC with CD1 mutant H65R. (A) Structure of P22-H65R BiFC constructs. P22 is a 20-amino-acid basic peptide derived from bacteriophage P22 that was fused to the N-terminus of H65R-NY and H65R-CY with a flexible hinge region between P22 and A3G. (B) Effect of P22 peptide on ability of H65R mutant to bind to RNA. 293T cells were co-transfected with pF-A3G and empty vector, or pF-A3G and A3G-CY, or pF-A3G and fourfold higher amount of P22-H65R-CY compared to pF-A3G. Co-IP assays were performed as described for Fig. 4A. (C) BiFC assays to evaluate interactions between wild-type and mutant A3Gs.