Clathrin-mediated endocytosis is modestly involved in the internalization of K_V10.1.

<p>A) Complexes of K_V10.1-BBS with BTX-Alexa594 (red) colocalize with clathrin-GFP (green) in punctuate structures (magnified insets, middle) compatible with endosomes. Line profile plots (bottom) through vesicles (white arrows) represent three classes of labeling. The colocalization map (right panel) highlights punctae with a high vs. low degree of colocalization. B) Surface-expression of K_V10.1-BBS is altered in cells that overexpress components of the clathrin-dependent endocytic machinery. Surface channels were labeled with BTX-biotin, isolated and immune-detected in western blots. Overexpression of the dominant-negative dynamin-K44A led to slight increases in surface expression, while overexpressing AP-180 slightly decreased K_V10.1-BBS surface-expression. C) K_V10.1 is internalized by fluid phase uptake. Complexes of K_V10.1-BBS with BTX-Alexa594 colocalize with dextran-rhodamine, a marker for fluid phase uptake, after 3 minutes of chase reaction. Corresponding ROIs from the merged dual-color image (center) and the intensity correlation image (right) highlight structures with colocalization.