Grossman Gregory H, Ebke Lindsey A, Beight Craig D, Jang Geeng-Fu, Crabb John W, Hagstrom Stephanie A
Department of Ophthalmic Research, Cleveland Clinic Cole Eye Institute, Cleveland, Ohio.
Vis Neurosci. 2013 Jul;30(4):129-39. doi: 10.1017/S0952523813000138. Epub 2013 Jun 10.
Dynamin proteins are involved in vesicle generation, providing mechanical force to excise newly formed vesicles from membranes of cellular compartments. In the brain, dynamin-1, dynamin-2, and dynamin-3 have been well studied; however, their function in the retina remains elusive. A retina-specific splice variant of dynamin-1 interacts with the photoreceptor-specific protein Tubby-like protein 1 (Tulp1), which when mutated causes an early onset form of autosomal recessive retinitis pigmentosa. Here, we investigated the role of the dynamins in the retina, using immunohistochemistry to localize dynamin-1, dynamin-2, and dynamin-3 and immunoprecipitation followed by mass spectrometry to explore dynamin-1 interacting proteins in mouse retina. Dynamin-2 is primarily confined to the inner segment compartment of photoreceptors, suggesting a role in outer segment protein transport. Dynamin-3 is present in the terminals of photoreceptors and dendrites of second-order neurons but is most pronounced in the inner plexiform layer where second-order neurons relay signals from photoreceptors. Dynamin-1 appears to be the dominant isoform in the retina and is present throughout the retina and in multiple compartments of the photoreceptor cell. This suggests that it may function in multiple cellular pathways. Surprisingly, dynamin-1 expression and localization did not appear to be disrupted in tulp1−/− mice. Immunoprecipitation experiments reveal that dynamin-1 associates primarily with proteins involved in cytoskeletal-based membrane dynamics. This finding is confirmed by western blot analysis. Results further implicate dynamin-1 in vesicular protein transport processes relevant to synaptic and post-Golgi pathways and indicate a possible role in photoreceptor stability.
发动蛋白参与囊泡的形成,为从细胞区室膜上切除新形成的囊泡提供机械力。在大脑中,发动蛋白-1、发动蛋白-2和发动蛋白-3已得到充分研究;然而,它们在视网膜中的功能仍不清楚。发动蛋白-1的一种视网膜特异性剪接变体与光感受器特异性蛋白类Tubby蛋白1(Tulp1)相互作用,Tulp1发生突变时会导致常染色体隐性遗传性视网膜色素变性的早发型。在此,我们利用免疫组织化学定位发动蛋白-1、发动蛋白-2和发动蛋白-3,并通过免疫沉淀结合质谱法探索小鼠视网膜中发动蛋白-1的相互作用蛋白,从而研究发动蛋白在视网膜中的作用。发动蛋白-2主要局限于光感受器的内节区室,提示其在外节蛋白运输中发挥作用。发动蛋白-3存在于光感受器的终末和二级神经元的树突中,但在内网状层最为明显,二级神经元在此中继来自光感受器的信号。发动蛋白-1似乎是视网膜中的主要异构体,存在于整个视网膜以及光感受器细胞的多个区室中。这表明它可能在多种细胞途径中发挥作用。令人惊讶的是,在tulp1−/−小鼠中,发动蛋白-1的表达和定位似乎并未受到破坏。免疫沉淀实验表明,发动蛋白-1主要与参与基于细胞骨架的膜动力学的蛋白相关联。蛋白质印迹分析证实了这一发现。结果进一步表明发动蛋白-1参与了与突触和高尔基体后途径相关的囊泡蛋白运输过程,并提示其在光感受器稳定性方面可能发挥作用。