TsCRISPR Based Identification of Rab Proteins Required for the Recycling of TRPL Ion Channel

Overview

Journal Front Cell Dev Biol

Specialty Cell Biology

Date 2024 Oct 7

PMID 39372952

Authors

Matthias Zeger

Lena Sarah Stanislawczyk

Marija Bulic

Andrea Maria Binder

Armin Huber

Affiliations

Soon will be listed here.

Abstract

In polarized cells, the precise regulation of protein transport to and from the plasma membrane is crucial to maintain cellular function. Dysregulation of intracellular protein transport in neurons can lead to neurodegenerative diseases such as Retinitis Pigmentosa, Alzheimer's and Parkinson's disease. Here we used the light-dependent transport of the TRPL (transient receptor potential-like) ion channel in photoreceptor cells to study the role of Rab proteins in TRPL recycling. TRPL is located in the rhabdomeric membrane of dark-adapted flies, but it is transported out of the rhabdomere upon light exposure and localizes at the Endoplasmatic Reticulum within 12 h. Upon subsequent dark adaptation, TRPL is recycled back to the rhabdomeric membrane within 90 min. To screen for Rab proteins involved in TRPL recycling, we established a tissue specific (ts) CRISPR/Cas9-mediated knock-out of individual genes in photoreceptors and assessed TRPL localization using an eGFP tagged TRPL protein in the intact eyes of these mutants. We observed severe TRPL recycling defects in the knockouts of , , , , . Using immunohistochemistry, we further showed that Rab3 and RabX2 each play a significant role in TRPL recycling and also influence TRPL transport. We localized Rab3 to the late endosome in photoreceptors and observed disruption of TRPL transport to the ER in knock-out mutants. TRPL transport from the ER to the rhabdomere ensues from the trans-Golgi where RabX2 is located. We observed accumulated TRPL at the trans-Golgi in knock-out mutants. In summary, our study reveals the requirement of specific Rab proteins for different steps of TRPL transport in photoreceptor cells and provides evidence for a unique retrograde recycling pathway of TRPL from the ER via the trans-Golgi.

Citing Articles

Comprehensive study of SNAREs involved in the post-Golgi transport in photoreceptors.

Ochi Y, Yamashita H, Sasaki S, Ogawa T, Yamada Y, Tago T Front Cell Dev Biol. 2024; 12:1442192.

PMID: 39720007 PMC: 11666571. DOI: 10.3389/fcell.2024.1442192.

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