Thompson Samantha L, Crowder Sophie M, Hekmatara Maryam, Sechrest Emily R, Deng Wen-Tao, Robichaux Michael A
Department of Ophthalmology & Visual Sciences and Department of Biochemistry & Molecular Medicine, West Virginia University, Morgantown, WV 26506, United States.
bioRxiv. 2024 Dec 16:2024.10.18.619115. doi: 10.1101/2024.10.18.619115.
Rod photoreceptor neurons in the retina detect scotopic light through the visual pigment rhodopsin (Rho) in their outer segments (OS). Efficient Rho trafficking to the OS through the inner rod compartments is critical for long-term rod health. Given the importance of protein trafficking to the OS, less is known about the trafficking of rod synaptic proteins. Furthermore, the subcellular impact of Rho mislocalization on rod synapses (i.e., "spherules") has not been investigated. In this study we used super-resolution and electron microscopies, along with proteomics, to perform a subcellular analysis of Rho synaptic mislocalization in P23H-Rho-RFP mutant mice. We discovered that mutant P23H-Rho-RFP protein mislocalized in distinct ER aggregations within the spherule cytoplasm, which we confirmed with AAV overexpression. Additionally, we found synaptic protein abundance differences in P23H-Rho-RFP mice. By comparison, Rho mislocalized along the spherule plasma membrane in WT and rd10 mutant rods, in which there was no synaptic protein disruption. Throughout the study, we also identified a network of ER membranes within WT rod presynaptic spherules. Together, our findings indicate that photoreceptor synaptic proteins are sensitive to ER dysregulation.
视网膜中的视杆光感受器神经元通过其外段（OS）中的视觉色素视紫红质（Rho）检测暗光。Rho通过视杆内段高效转运至OS对于视杆细胞的长期健康至关重要。鉴于蛋白质向OS转运的重要性，人们对视杆突触蛋白的转运了解较少。此外，Rho定位错误对视杆突触（即“小球”）的亚细胞影响尚未得到研究。在本研究中，我们使用超分辨率显微镜和电子显微镜以及蛋白质组学，对P23H-Rho-RFP突变小鼠中Rho突触定位错误进行亚细胞分析。我们发现突变型P23H-Rho-RFP蛋白在小球细胞质内不同的内质网聚集体中定位错误，我们通过腺相关病毒过表达证实了这一点。此外，我们发现P23H-Rho-RFP小鼠中突触蛋白丰度存在差异。相比之下，在野生型和rd10突变型视杆中，Rho沿小球质膜定位错误，其中突触蛋白未受破坏。在整个研究过程中，我们还在野生型视杆突触前小球内鉴定出一个内质网膜网络。总之，我们的研究结果表明光感受器突触蛋白对内质网失调敏感。
