Fig. 2. Turnover of full-length APP and APP-αCTF in LRP–/– cells. LRP+/– and LRP–/– fibroblasts were pulse labeled with [³⁵S]methionine/cysteine for 15 min and chased for 0, 1, 2 and 4 h (A). At time 0, APP consists predominantly of immature N-glycosylated species. Both the N-glycosylated and mature N- and O-glycosylated species are abundant at 1 h for both cell types. After a 2 h chase period, the APP level is dramatically reduced in LRP+/– cells and hardly detectable at 4 h. In contrast, APP can still be detected in LRP–/– cells even after a 4 h chase period. (B) The half-life was determined by quantitating the results from (A) as shown. (C) APP-αCTF turnover was determined by metabolically labeling LRP+/– and LRP–/– fibroblasts with [³⁵S]methionine/cysteine for 1 h and chasing for 3, 6 and 18 h. Similar amounts of APP-αCTFs are present after the labeling and after the 3 h chase period in LRP+/– and LRP–/– cells. Even after an 18 h chase period, APP-αCTFs can still be detected in LRP+/– fibroblasts. However, APP-αCTFs in LRP–/– fibroblasts were almost completely degraded. (D) The half-life of APP-αCTF was determined by quantitating the results from (C) as shown. The experiment in (A) was performed in triplicate and that in (C) in duplicate, and representative experiments are shown.