Figure 2
Juvenile hemochromatosis-linked RGMc COOH-terminal truncation mutants are cleaved into smaller isoforms. A. Map of mouse RGMc showing locations of signal peptide (s.p.), RGD sequence, von Willebrandt type D domain, GPI-anchor motif, and putative PC site (R-X-X-R). RGMcΔGPI-His and RGMc frame-shift truncation mutants are pictured below. Each contains a COOH-terminal His tag. B. Detection of RGMc truncation mutants by immunoblotting of protein lysates and conditioned medium from transiently transfected Cos-7 cells using RGMc (upper panel) or HA antibodies (lower). C. Detection of RGMc truncation mutants by immunoblotting of conditioned medium after incubation ± RVKR using RGMc (upper panel) or His antibodies (lower). For B and C, black arrows indicate 50 kDa single-chain RGMc and white arrows 40 kDa, and gray arrows mark NH2- and COOH-terminal fragments resulting from intra-molecular cleavage.