Figure 4
Furin cleaves soluble RGMc to produce the 40 kDa isoform. A. Only 50 kDa RGMc accumulates in culture medium of cells expressing a protein with a modified PC cleavage site. Immunoblot of soluble RGMc from conditioned medium of Cos-7 cells transfected with either wild-type (WT) RGMc or a derivative containing a mutated PC site (RXXRmut), after incubation ± RVKR. Similar results were observed with Hep3B cells. B. Hep3B cells cleave 50 kDa RGMc. RGMc was added to medium of Hep3B cells for 24 h ± RVKR, followed by immunoblotting (nt = non-treated). Identical results were seen with Cos-7 cells. C. Furin cleaves 50 kDa RGMc. Immunoblot shows results of in vitro incubation of soluble 50 kDa HA-RGMc ± recombinant furin. Antibodies are indicated. For A – C, arrows are as in legend to Fig. 1.