TGase2 interacts with TfR1 and enhances mesangial IgA1 deposition. (A) Binding of TGase2 to different types of IgA detected by ELISA using biotinylated TGase2. Albumin was used as a negative control. n = 3 experiments. (B–D) Double immunofluorescence for human IgA and TGase2 (B), fibronectin and TGase2 (labeled with anti–rabbit Alexa Fluor 568; C), and fibronectin and human IgA1 (D) in frozen sections from α1KI and α1KI-CD89Tg mice. Bars, 10 µm. (E) Binding of TGase2 to TfR1-His detected by ELISA using biotinylated TGase2. β-Synuclein (His-tagged protein) was used as a negative control. n = 4 experiments. (A and E) Error bars indicate SEM. (F) TGase2 was loaded on a column of nickel beads preincubated or not with sTfR1-His. Eluates were analyzed by SDS–10% PAGE followed by Western blot using an anti-TGase2 antibody plus anti–rabbit IgG-HRP. (G) Flow cytometry histogram (one representative experiment of six) of HMCs binding TGase2 (white layer) using biotinylated-TGase2 plus streptavidin-APC. The dashed layer represents incubation with sTfR1. The gray layer corresponds to HMCs with streptavidin-APC.