FIGURE 6.

hENT3 is predominantly degraded by the lysosomal pathway. NIH 3T3 fibroblasts expressing hENT3HA were treated with dimethyl sulfoxide (control), 20 μm MG132 (proteasomal inhibitor), or 10 μm leupeptin (lysosomal inhibitor) for 16 h, and hENT3 was accumulation examined. A, immunostaining analysis of hENT3 (red) using 1:1000 rabbit anti-HA polyclonal antibody indicated increased staining intensity of hENT3 in leupeptin-treated cells (bottom panel) but not in MG132-treated cells (middle panel). The original magnification is ×40. B, Western blotting analysis indicated a higher accumulation of hENT3 (∼65 kDa) in leupeptin-treated cells (right lane) than seen in MG132-treated cells (middle lane). β-Actin (45 kDa) was used as a loading control.