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. 2010 Aug;24(8):2783–2794. doi: 10.1096/fj.09-146357

Figure 5.

Figure 5

Reducing SNX6 increases basal BACE1 levels. A) Reducing SNX6 increases endogenous BACE1 levels. Neuro2a cells stably expressing the Swedish variant of APP were transfected with either nonsilencing shRNA vector (NS) or shRNA against mouse SNX6 (SNX6). After 2 d incubation, cell lysates were subjected to Western blot analysis. B) Quantification of BACE1 was performed using NIH ImageJ (n=3). Values are means ± sd. *P < 0.005; 2-tailed t test. C) Reducing SNX6 does not affect BACE1 mRNA levels. HeLa cells were transfected with 2 distinct SNX6 shRNAs (SNX6 RNAi) or negative control shRNAs (NS). Real-time PCR was performed, and mRNA levels of SNX6 and BACE1, normalized to β-actin mRNA, were quantified. D) Pulse-chase experiments using Halo-tagged BACE1. HEK293 cell lines stably expressing either nonsilencing shRNA vector (NS) or shRNA against human SNX6 (SNX6; see Fig. 4A) were transfected with Halo-tagged BACE1. After 1 d of incubation, cells were labeled with biotin-conjugated Halo-tag ligand and chased for indicated times. BACE1 turnover was analyzed with Western blot analysis. Representative blots are shown. E) Protein levels of BACE1 were plotted with the chased times. Error bars = sd from 3 experiments.