Signals Governing the Trafficking and Mistrafficking of a Ciliary GPCR, Rhodopsin
Signals Governing the Trafficking and Mistrafficking of a Ciliary GPCR, Rhodopsin
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DOI：
10.1523/jneurosci.1520-13.2013
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发表时间：
2013-08-21
影响因子：
5.3
通讯作者：
Imanishi, Yoshikazu
中科院分区：
文献类型：
--
作者：
Lodowski, Kerrie H.;Lee, Richard;Imanishi, Yoshikazu
视紫红质是一种对视觉至关重要的纤毛特异性GPCR。视紫红质定位错误与称为视网膜纤毛病变的致盲疾病有关。视紫红质错误定位于视杆细胞感光神经元的机制还不清楚。因此，我们研究了运输信号在视紫红质错误定位中的作用。将视紫红质及其截短突变体与光转换荧光蛋白Dendra 2融合，并在非洲爪蟾视杆细胞中表达。Dendra 2的光转化导致颜色从绿色变为红色，使得能够可视化与视紫红质运输和更新相关的动态事件。我们发现视紫红质的错误定位是一个易化的过程，其中位于322-326 aa（CCGKN）内的信号是必不可少的。327-336 aa内的额外信号进一步促进了错误定位。当VXPX纤毛靶向基序不存在时，这种集体错误信号会对视紫红质产生毒性并导致错误定位。我们还确定，VXPX基序中和这种mistrafficking信号，增强纤毛靶向至少10倍，并加速后高尔基体囊泡结构的贩运。在缺乏VXPX基序的情况下，错误定位的视紫红质通过囊泡分泌到细胞外环境中而被主动清除。因此，本研究揭示了运输信号在视紫红质定位和更新中的多重作用。
Rhodopsin is a cilia-specific GPCR essential for vision. Rhodopsin mislocalization is associated with blinding diseases called retinal ciliopathies. The mechanism by which rhodopsin mislocalizes in rod photoreceptor neurons is not well understood. Therefore, we investigated the roles of trafficking signals in rhodopsin mislocalization. Rhodopsin and its truncation mutants were fused to a photo-convertible fluorescent protein, Dendra2, and expressed in Xenopus laevis rod photoreceptors. Photoconversion of Dendra2 causes a color change from green to red, enabling visualization of the dynamic events associated with rhodopsin trafficking and renewal. We found that rhodopsin mislocalization is a facilitated process for which a signal located within 322-326 aa (CCGKN) is essential. An additional signal within 327-336 aa further facilitated the mislocalization. This collective mistrafficking signal confers toxicity to rhodopsin and causes mislocalization when the VXPX cilia-targeting motif is absent. We also determined that the VXPX motif neutralizes this mistrafficking signal, enhances ciliary targeting at least 10-fold, and accelerates trafficking of post-Golgi vesicular structures. In the absence of the VXPX motif, mislocalized rhodopsin is actively cleared through secretion of vesicles into the extracellular milieu. Therefore, this study unveiled the multiple roles of trafficking signals in rhodopsin localization and renewal.