Targeted delivery of rhodopsin's assembled core is required for outer segment extension in mouse rod photoreceptors

Vertebrate vision in dim-light environments is initiated by rod photoreceptor cells that express the photopigment rhodopsin, a G-protein coupled receptor (GPCR). To ensure efficient light capture, rhodopsin is densely packed into hundreds of membrane discs that are tightly stacked within the rod-shaped outer segment compartment. Along with its role in eliciting the visual response, rhodopsin serves as both a building block necessary for proper outer segment formation as well as a trafficking guide for a few outer segment resident membrane proteins. An interesting aspect of rod homeostasis is that mutations that affect the localization of rhodopsin to the outer segment result in photoreceptor degeneration. In this study, we focus on determining the necessary properties of rhodopsins cytosolic C-terminus required for either proper outer segment trafficking and/or the capacity to extend the rudimentary outer segment in rhodopsin knockout (RhoKO) rods. We find that the well-described C-terminal QVAPA outer segment targeting motif also plays a role in endoplasmic reticulum (ER) exit and is necessary for outer segment elongation. Even minor rhodopsin mislocalization prevents efficient outer segment elongation suggesting that QVAPA-targeted delivery of rhodopsin drives outer segment extension. We identify that rhodopsins core, helix-8, and accessible QVAPA targeting motif are the minimal requirements to extend the rudimentary outer segment in RhoKO rods. Our findings provide useful insights into rhodopsins molecular features needed for outer segment delivery and subsequent elongation of this membrane-rich compartment.