Retinal Ciliopathy gene SPATA7, plays a critical role in maintaining the integrity of Transition zone complex in photoreceptors

Abstract

Purpose: SPATA7 was previously identified by our group as a causal gene for Leber congenital amaurosis (LCA) and Juvenile retinitis pigmentosa (RP), loss of which leads to Rhodopsin mis-localization and rapid photoreceptor degeneration in mice. We have recently established that SPATA7 localizes at the connecting cilium (CC) and interacts with RPGRIP1, a crucial protein involved in protein trafficking at the transition zone (TZ). The purpose of this study is to further our understanding of SPATA7 by identifying its additional interacting proteins, which would shed light on its function in the photoreceptor.

Methods: Both IP-MS from various mouse tissues using Wild type, Spata7-GFP, Spata7-Flag transgenic mice lines as well as in vitro protein interaction screen have been performed. The interactions were confirmed by performing in vivo immunoblotting analysis on retina tissue.

Results: SPATA7 interacts with a number of CC proteins such as INPP5E, SDCCAG8, ATXN10, NPHP1, which are critical protein components of the RPGRIP1 transition zone complex. These interactions are functionally relevant as SPATA7 localizes to the CC photoreceptors and is an interacting partner of RPGRIP1. Immunoblotting using extracts of 293T cells confirms interaction of INPP5E, SDCCAG8 with SPATA7. Strikingly, analysis of the OS proteome suggests that subcellular localization of these interacting proteins is affected in the Spata7 KO mice mutant retina.

Conclusions: Our research suggests that SPATA7 is a crucial member of the TZ complex in the absence of which crucial TZ proteins are mis-localized. Based on these results, we propose a model where SPATA7 acts as a scaffold to these TZ proteins, maintaining the integrity of the TZ complex and thereby stabilizing the cilia transport.