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. 2008 Aug 1;283(31):21427–21432. doi: 10.1074/jbc.M800014200

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Copyright © 2008, The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 2. — DKK1 does not affect the cell surface LRP6 level. A, DKK1 does not affect the cell surface LRP6 level in L (panel a), Rat2 (panel b), and MEF (panel c) cells. After cells were treated with DKK1 CM for the indicated period of time, cell surface proteins were biotinylated, extracted and precipitated with avidin-agarose beads, and analyzed for the presence of LRP6 protein by immunoblotting. B, DKK1 does not affect the cell surface or total LRP6 level in HEK293T cells. HEK293T cells were untreated (lane 1) or treated with DKK1 CM for 2 h (lane 2), and cell surface proteins were biotinylated, extracted, and precipitated with LRP6 antibodies. The amount of LRP6 on the cell surface was determined via immunoblotting with avidin-HRP (panel a), and the total amount of LRP6 in precipitates was determined with the LRP6 antibodies (panel b). C, EGF induces rapid down-regulation of EGFR on the cell surface. Rat2 cells were untreated (lane 1) or treated with 100 ng/ml EGF for 15 min (lane 2), and cell surface proteins were biotinylated, extracted, precipitated with avidin-agarose beads, and analyzed for the presence of EGFR (panel a) or LRP6 (panel b) by immunoblotting. Cell extracts were also analyzed for the total EGFR (panel c) and LRP6 (panel d) proteins.