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. 2012 Apr 1;23(7):1294–1306. doi: 10.1091/mbc.E11-09-0780

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© 2012 Ishikawa et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 4: — LRRK1(Y944F) leads to EGF accumulation in endosomal compartments in the perinuclear region. (A–D) Distribution of Rh-EGF. HeLa S3 cells were transfected with wild-type GFP-LRRK1 (A), GFP-LRRK1(Y944F) (B, D), and GFP-LRRK1(Y944F; K1243M) (C), as indicated. After 16 h of serum starvation, cells were briefly stimulated with or without Rh-EGF (40 ng/ml), followed by washing to remove labeled EGF from the medium. The cells shown in D were preincubated with nocodazole (5 μg/ml) for 30 min before EGF stimulation. Cells were incubated for the indicated times after the initial exposure to Rh-EGF and then fixed and stained with 4′,6-diamidino-2-phenylindole. Yellow colors in the merged images indicate colocalization of GFP-LRRK1 and Rh-EGF. Scale bar, 10 μm. (E) Quantification of the EGF accumulation in the perinuclear region. Histogram indicates the percentage of cells that have endosomes (>2.0 μm diameter) containing Rh-EGF in the perinuclear region. Values reflect the mean SD of three independent experiments, with an average of 50 cells scored per samples. Data are compared using a two-tailed unpaired Student's t test. *p < 0.05; **p < 0.01; NS, not significant.