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. 2017 Oct 2;216(10):3323–3338. doi: 10.1083/jcb.201701034

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© 2017 Fiuza et al.

This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).

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Figure 3. — PICK1 is recruited to CCPs in an AP2-dependent and a dynamin-independent manner in dynamin TKO fibroblasts. (A) PICK1 is localized on arrested CCP necks labeled with endophilin-A2. Dynamin TKO fibroblasts expressing GFP-WT-PICK1 and endophilin-A2–mCherry were imaged live by spinning-disk confocal microscopy. A’ shows magnification from boxed region. (A, right) Kymograph shows that the PICK1–endophilin-A2 colocalization is stable for several minutes. (Note that these cells accumulate arrested CCPs.) (B) PICK1 colocalizes with the clathrin coat. Dynamin TKO fibroblasts expressing GFP-WT-PICK1 and clathrin LC–mRFP were imaged live by spinning-disk confocal microscopy. B’ shows magnification from boxed region. (B, right) Kymograph shows that the PICK1–clathrin LC (CLC) colocalization is stable for several minutes. (C) Localization of PICK1 to CCPs requires interaction with AP2. Dynamin TKO fibroblasts expressing GFP-DDAA-PICK1 and clathrin LC–mRFP were imaged live by confocal microscopy. C’ shows magnification from boxed region. (C, right) Kymograph shows that GFP-DDAA-PICK1 does not colocalize with clathrin LC over time. Bars: (whole cells) 10 µm; (magnified images) 5 µm.