Local trafficking at dendritic branch points. (A) A cluster of VSVG-mEOS2-2×UVR8 was photoswitched (arrow marked 405 nm) in a dendrite before global UV-B treatment (arrow marked UV-B) to release labeled cargo from the ER. Note that VSVG-mEOS2-2×UVR8 clusters remain intact after focal 405-nm illumination and can be subsequently dissolved with UV-B. Bar, 1 µm. (B) Experimental strategy. Branch point cargo can be selectively tagged and tracked by photoconverting mEOS-tagged VSVG-2×UVR8 before ER release with UV-B. (C) Branch point cargo was selectively photoconverted to red with local 405-nm illumination (crosshair, middle) and then released with global UV-B. Note the accumulation of red puncta at sites near the branch point after UV-B treatment (arrowheads). Not all nearby puncta contained photoswitched cargo (asterisks), indicating selective trafficking to a subset of branch point organelles. Bar, 4 µm. (D) Tracking non-branch point ER cargo. No local accumulation of secretory cargo was observed when we tagged ER cargo at sites distant from branch points. Bar, 3 µm. (E) Quantification of local photoswitching branch points vs. non-branch point ER before release. Branch point cargo was redistributed, but effectively retained for tens of minutes near its origin while non-branch point cargo quickly diffused away and diluted into the bulk, nonphotoconverted VSVG. n = at least 3 branch points and 5 different non-branch point sites from 3 different neurons. Data collected from three independent experiments. The shaded regions in the traces indicate the range of standard error of the mean.