Figure 7.

Loss of sorting nexin 9 results in increased shedding of an ADAM9 substrate. A and B, MDA-MB-231 cells were transfected with ADAM9 or control siRNAs, and subsequently with the ADAM9 substrate, ephrin B4 receptor N-terminally fused to alkaline phosphatase (EphB4-AP). Cells were incubated for 18 h, and in A, cell lysates were analyzed by immunoblotting (IB) as indicated, or B, the amount of EphB4-AP shed into the conditioned cell media was measured, n = 3. C and D, HEK293-VnR cells were co-transfected with EphB4-AP and WT GFP-tagged ADAM9 (ADAM9-WT-GFP), catalytic inactive GFP-tagged ADAM9 (ADAM9-EA-GFP), or control vector (EV-GFP). Cells were incubated for 18 h, and cell lysates were analyzed as in A and B, n = 3. E and F, HEK293-VnR cells were transfected with SNX9 or control siRNA and subsequently co-transfected with EphB4-AP. Cells were incubated for 18 h with or without Batimastat, and cell lysates were analyzed as in A and B, n = 3. Western blots are representative of n = 3 independent experiments. Plots show individual data, and average values ± S.D., *, p < 0.05 (ANOVA).