Restoration of the majority of the visual spectrum by using modified Volvox channelrhodopsin-1.
Restoration of the majority of the visual spectrum by using modified Volvox channelrhodopsin-1.
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DOI：
10.1038/mt.2014.81
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发表时间：
2014-08
期刊：
影响因子：
--
通讯作者：
Tamai M
中科院分区：
文献类型：
--
作者：
Tomita H;Sugano E;Murayama N;Ozaki T;Nishiyama F;Tabata K;Takahashi M;Saito T;Tamai M
我们以前表明，失明的大鼠的视力恢复衣原体通道视紫红质-2（ChR 2）的基因转移只能检测波长小于540 nm，因为转基因产品的作用光谱。团藻衍生的通道视紫红质-1，VChR 1，具有比ChR 2更宽的光谱。然而，VChR 1蛋白主要定位于细胞质中，并显示出弱的离子通道特性时，VChR 1基因转染到HEK 293细胞。我们产生了改性团藻通道视紫红质-1（mVChR 1），这是团藻通道视紫红质-1和衣原体通道视紫红质-1的嵌合体，并表现出增加质膜整合和显着改善其通道特性。在全细胞膜片钳下，mVChR 1表达细胞在468-640 nm刺激时显示光诱导电流。mVChR 1表达细胞中的诱发电流比VChR 1表达细胞中的诱发电流大约30倍。从遗传学上讲，通过腺相关病毒载体表达mVChR 1的失明大鼠恢复了对波长在468和640 nm之间的光的视觉反应，并且恢复的视觉反应维持了一年。因此，mVChR 1是用于恢复视力的基因治疗的候选基因，并且mVChR 1的基因递送可以为盲人患者提供大部分可见光谱。
We previously showed that blind rats whose vision was restored by gene transfer of Chlamydomonas channelrhodopsin-2 (ChR2) could only detect wavelengths less than 540 nm because of the action spectrum of the transgene product. Volvox-derived channelrhodopsin-1, VChR1, has a broader spectrum than ChR2. However, the VChR1 protein was mainly localized in the cytoplasm and showed weak ion channel properties when the VChR1 gene was transfected into HEK293 cells. We generated modified Volvox channelrhodopsin-1 (mVChR1), which is a chimera of Volvox channelrhodopsin-1 and Chlamydomonas channelrhodopsin-1 and demonstrated increased plasma membrane integration and dramatic improvement in its channel properties. Under whole-cell patch clamp, mVChR1-expressing cells showed a photo-induced current upon stimulation at 468–640 nm. The evoked currents in mVChR1-expressing cells were ~30 times larger than those in VChR1-expressing cells. Genetically, blind rats expressing mVChR1 via an adeno-associated virus vector regained their visual responses to light with wavelengths between 468 and 640 nm and their recovered visual responses were maintained for a year. Thus, mVChR1 is a candidate gene for gene therapy for restoring vision, and gene delivery of mVChR1 may provide blind patients access to the majority of the visible light spectrum.
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