833A Poster - 13. Neural development and physiology
Thursday April 07, 2:00 PM - 4:00 PM

Authors:
Josephine Mitchell ¹; Ipek Midillioglu ²; Jill Wildonger ^2,4; Kathy Wang ³; Chun Han ³

Affiliations:
1) Biochemistry Department, University of Wisconsin-Madison, Madison, WI; 2) Pediatrics Department, University of California, San Diego, La Jolla, CA; 3) Weill Institute for Cell and Molecular Biology and Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY; 4) Division of Biological Sciences, Cell & Developmental Biology Section, University of California, San Diego, La Jolla, CA

Keywords:
j. ion channels; e. intracellular transport

An organism’s ability to perceive stimuli, such as pain, from its surrounding environment is essential to survival. The perception of an external stimuli and its transformation into a cellular response depends on ion channels that are distributed throughout the dendritic arbor of peripheral sensory neurons. However, it is not well understood how trafficking of ion channels to their sites of function in the dendritic membrane is regulated and whether or how their
localization is coordinated with dendrite morphogenesis. We investigated how ion channel localization is regulated using the Drosophila melanogaster class IV dendritic arborization neurons. In larvae, painful mechanical stimuli are sensed in part by pickpocket 1 (ppk1) and pickpocket 26 (ppk26), which are members of the degenerin/epithelial Na+ channel/acid sensing ion channel (DEG/ENaC/ASIC) family. Ppk1 and ppk26 form a heterotrimeric channel in
the class IV sensory dendrites and are dependent on each other for membrane insertion. To study the regulation of pickpocket ion channel trafficking, we used CRISPR-Cas9 genome engineering to fluorescently tag endogenous ppk1. We found that ppk1 localizes robustly and uniformly throughout the dendritic membrane and is present in actively growing dendrite tips, indicating that sensory function is established during dendrite morphogenesis. To delineate mechanisms that regulate ppk1 trafficking we investigated the endocytic network. We hypothesized that Rab11 may be involved in forward trafficking of ppk channels to the membrane and/or through local regulation via recycling pathways. We found that ppk1 and ppk26 localization to the dendritic membrane depends on the recycling endosome GTPase Rab11. Additionally, we found that ppk1 and ppk26 colocalize with the early endosome GTPase Rab5. When ppk1 is absent, ppk26 is not membrane expressed and instead forms puncta throughout the dendrites. Over half of these ppk26 puncta colocalize with Rab5, suggesting involvement of early endosomes in quality control of pickpocket channel trafficking. Together, our results suggest that pickpocket channel trafficking is coordinated with sensory dendrite morphogenesis and that its membrane expression relies on the endosomal network.