Grayson Celene, Bartolini Francesca, Chapple J Paul, Willison Keith R, Bhamidipati Arunashree, Lewis Sally A, Luthert Philip J, Hardcastle Alison J, Cowan Nicholas J, Cheetham Michael E
Division of Pathology, Institute of Ophthalmology, UCL, London EC1V 9EL, UK.
Hum Mol Genet. 2002 Nov 15;11(24):3065-74. doi: 10.1093/hmg/11.24.3065.
Mutations in the retinitis pigmentosa 2 (RP2) gene cause a severe form of X-linked retinal degeneration. RP2 is a ubiquitous 350 amino acid plasma membrane-associated protein, which shares homology with the tubulin-specific chaperone cofactor C. RP2 protein, like cofactor C, stimulates the GTPase activity of tubulin in combination with cofactor D. RP2 has also been shown to interact with ADP ribosylation factor-like 3 (Arl3) in a nucleotide and myristoylation-dependant manner. In this study we have examined the relationship between RP2, cofactor C and Arl3 in patient-derived cell lines and in the retina. Examination of lymphoblastoid cells from patients with an Arg120stop nonsense mutation in RP2 revealed that the expression levels of cofactor C and Arl3 were not affected by the absence of RP2. In human retina, RP2 was localized to the plasma membrane of cells throughout the retina. RP2 was present at the plasma membrane in both rod and cone photoreceptors, extending from the outer segment through the inner segment to the synaptic terminals. There was no enrichment of RP2 staining in any photoreceptor organelle. In contrast, cofactor C and Arl3 localized predominantly to the photoreceptor connecting cilium in rod and cone photoreceptors. Cofactor C was cytoplasmic in distribution, whereas Arl3 localized to other microtubule structures within all cells. Arl3 behaved as a microtubule-associated protein: it co-localized with microtubules in HeLa cells and this was enhanced following microtubule stabilization with taxol. Furthermore, Arl3 co-purified with microtubules from bovine brain. Following microtubule depolymerization with nocodazole, Arl3 relocalized to the nuclear membrane. These data suggest that RP2 functions in concert with Arl3 to link the cell membrane with the cytoskeleton in photoreceptors as part of the cell signaling or vesicular transport machinery.
视网膜色素变性2（RP2）基因的突变会导致一种严重的X连锁视网膜变性。RP2是一种普遍存在的含350个氨基酸的质膜相关蛋白，与微管蛋白特异性伴侣辅助因子C具有同源性。与辅助因子C一样，RP2蛋白与辅助因子D共同刺激微管蛋白的GTPase活性。RP2还被证明以核苷酸和肉豆蔻酰化依赖的方式与ADP核糖基化因子样3（Arl3）相互作用。在本研究中，我们研究了患者来源的细胞系和视网膜中RP2、辅助因子C和Arl3之间的关系。对RP2基因发生Arg120stop无义突变患者的淋巴母细胞进行检测发现，辅助因子C和Arl3的表达水平不受RP2缺失的影响。在人类视网膜中，RP2定位于整个视网膜细胞的质膜。RP2存在于视杆和视锥光感受器的质膜中，从外段延伸至内段再到突触终末。在任何光感受器细胞器中均未发现RP2染色增强。相比之下，辅助因子C和Arl3主要定位于视杆和视锥光感受器的光感受器连接纤毛。辅助因子C分布于细胞质中，而Arl3定位于所有细胞内的其他微管结构。Arl3表现为一种微管相关蛋白：它在HeLa细胞中与微管共定位，在用紫杉醇稳定微管后这种共定位增强。此外，Arl3与来自牛脑的微管共纯化。用诺考达唑使微管解聚后，Arl3重新定位于核膜。这些数据表明，RP2与Arl3协同作用，作为细胞信号传导或囊泡运输机制的一部分，将光感受器中的细胞膜与细胞骨架连接起来。
