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. Author manuscript; available in PMC: 2019 Apr 3.
Published in final edited form as: Cell Metab. 2018 Mar 22;27(4):816–827.e4. doi: 10.1016/j.cmet.2018.02.010

Figure 5. γ-secretase regulates endocytosis of TRLs.

Figure 5

(A and B) Western blot of hApoB and hApoC3 in media of primary hepatocytes from Cre- or L-Ncst mice (A) and shControl or shNcst-stable McA-RH7777 cells (B), with Pitstop 2. (C and D) Western blot of hepatic LDLR in Cre- or L-Ncst (C), or shControl or shNcst cells (D). (E) Cell surface biotinylation analysis in shControl or shNcst-stable McA-RH7777 cells. (F) Western blot of exogenous LDLR with C-terminal Flag tags in shControl or shNcst-stable McA-RH7777 cells. (G, H) Western blots for C-terminal GFP-tagged LDLR (or α-tubulin), in the presence of epoxomicin and/or GSI. FL=full-length LDLR, CTD=LDLR C-terminal domain; *= nonspecific band, **=C-terminal LDLR fragment. (I) Western blots of co-immunoprecipitated Nicastrin, in McA-RH7777 cells transfected with Flag-tagged full-length (FL) or LDLR lacking the C-terminal domain (dCTD). (J) Western blots from McA-RH7777 cells transfected with Flag-tagged FL or dCTD, then treated with 50 μg ml−1 cycloheximide (CHX) for the indicated times, and quantification of Flag normalized to β-actin, relative to time 0. (K) McA-RH7777 cells transfected with LDLR-dCTD-Flag plasmids, pretreated with chloroquine for 1 h before incubation with CHX for 4 hrs. (L) Western blots from shControl or shNcst-stable McA-RH7777 cells, transfected with FL- or LDLR-dCTD-Flag. *P < 0.05 as compared to the indicated control by two-way ANOVA. All data are shown as the means ± s.e.m.