Departments of Ophthalmology.
Departments of Ophthalmology,
J Neurosci. 2019 May 1;39(18):3376-3393. doi: 10.1523/JNEUROSCI.2811-18.2019. Epub 2019 Feb 28.
Peripherin 2 (PRPH2) is a tetraspanin protein concentrated in the light-sensing cilium (called the outer segment) of the vertebrate photoreceptor. The mechanism underlying the ciliary targeting of PRPH2 and the etiology of cone dystrophy caused by PRPH2 mutations remain elusive. Here we show that the late endosome (LE) is the main waystation that critically sorts newly synthesized PRPH2 to the cilium. PRPH2 is expressed in the luminal membrane of the LE. We delineate multiple C-terminal motifs of PRPH2 that distinctively regulate its LE and ciliary targeting through ubiquitination and binding to ESCRT (Endosomal Sorting Complexes Required for Transport) component Hrs. Using the newly developed TetOn-inducible system in transfected male and female mouse cones , we show that the entry of nascent PRPH2 into the cone outer segment can be blocked by either cone dystrophy-causing C-terminal mutations of PRPH2, or by short-term perturbation of the LE or recycling endosomal traffic. These findings open new avenues of research to explore the biological role of the LE in the biosynthetic pathway and the etiology of cone dystrophy caused by PRPH2 mutations and/or malfunctions of the LE. Peripherin 2 (PRPH2) is a tetraspanin protein abundantly expressed in the light-sensing cilium, the outer segment, of the vertebrate photoreceptor. The mechanism underlying the ciliary transport of PRPH2 is unclear. The present study reveals a novel ciliary targeting pathway, in which the newly synthesized PRPH2 is first targeted to the lumen of the late endosome (LE) en route to the cilia. We deciphered the protein motifs and the machinery that regulates the LE trafficking of PRPH2. Using a novel TetOn-inducible system in transfected mouse cones, we showed that the LE pathway of PRPH2 is critical for its outer segment expression. A cone dystrophy-causing mutation impairs the LE and ciliary targeting of PRPH2, implicating the relevance of LE to cone/macular degenerative diseases.
外周蛋白 2 (PRPH2) 是一种四跨膜蛋白，在脊椎动物感光器的光感受器纤毛（称为外节段）中高度表达。PRPH2 向纤毛的睫状靶向的机制和由 PRPH2 突变引起的锥细胞营养不良的病因仍然难以捉摸。在这里，我们表明晚期内体 (LE) 是将新合成的 PRPH2 分类到纤毛的主要中转站。PRPH2 在外体的腔膜上表达。我们描绘了 PRPH2 的多个 C 末端基序，这些基序通过泛素化和与内体分选复合物必需运输 (ESCRT) 成分 Hrs 结合，分别调节其 LE 和纤毛靶向。使用新开发的 TetOn 诱导系统在转染的雄性和雌性小鼠锥体中，我们表明新生 PRPH2 进入锥体外节段的进入可以被 PRPH2 的尾部引起的锥体营养不良突变或 LE 和循环内体运输的短期扰动所阻断。这些发现为探索 LE 在生物合成途径中的生物学作用以及由 PRPH2 突变和/或 LE 功能障碍引起的锥细胞营养不良的病因开辟了新的研究途径。外周蛋白 2 (PRPH2) 是一种四跨膜蛋白，在脊椎动物感光器的光感受器纤毛（称为外节段）中大量表达。PRPH2 向纤毛的运输机制尚不清楚。本研究揭示了一种新的纤毛靶向途径，其中新合成的 PRPH2 首先靶向晚期内体 (LE) 的腔，然后再靶向纤毛。我们破译了调节 PRPH2 LE 运输的蛋白质基序和机制。使用转染的小鼠锥体中的新型 TetOn 诱导系统，我们表明 PRPH2 的 LE 途径对其外节段表达至关重要。导致锥细胞营养不良的突变会损害 PRPH2 的 LE 和纤毛靶向，这表明 LE 与锥细胞/黄斑退行性疾病有关。
