Figure 5.

Defective cellular trafficking causes loss of collagen-induced signaling for the SMED-SL patient mutant variants T713I-DDR2, I726R-DDR2 and R752C-DDR2. HA-tagged full-length DDR2 wild-type or mutant variants were transiently expressed in HEK293 cells. (A) Cell lysates were treated with Endoglycosidase H for 3 h at 37°C (H) or left untreated for 3 h at 37°C (−) and analyzed by SDS–PAGE and western blotting. The blot was probed with polyclonal anti-DDR2 antibodies. (B) Cells were stimulated with 10 µg/ml of rat tail collagen I (+) or 1 mm acetic acid (−) for 90 min at 37°C. Cell lysates were analysed by SDS–PAGE and western blotting. The blots were probed with anti-phosphotyrosine (anti-PY) monoclonal antibody 4G10 (upper blot) or polyclonal anti-DDR2 antibodies (lower blot). The positions of molecular markers (in kDa) are indicated. The experiments were carried out three times with very similar results.