Functional and Genomic Study of the Novel (Orphan) Opsin Profile of the Zebrafish

视网膜是一层薄薄的相互连接的神经细胞，覆盖在眼球后部的内表面。视觉本身的整个过程始于一小群蛋白质（称为视蛋白），它们执行吸收光并将其转化为电信号/生物信号的独特任务。在视网膜中，传统上认为这些视蛋白局限于视杆细胞和视锥细胞，统称为光感受器。这些感光细胞的功能相当于数码相机CCD阵列中的像素。最近已经发现，实际上在视网膜的替代细胞中产生了丰富多样的额外候选视蛋白，但它们的功能在很大程度上仍未被探索。此外，在大脑的松果体器官等部位也发现了同样的光敏蛋白。总的来说，这些观测表明，光探测的整个过程比原先想象的要复杂得多。在当前的项目中，我们建议对这些潜在的光感应蛋白进行系统的研究，通过观察斑马鱼（一种常见的模式生物）中整个不同的视蛋白光色素家族。最初，我们将把编码这些蛋白质的DNA插入到培养条件下的宿主细胞中。通过这种方式，我们可以测量和比较它们的功能特性，并研究它们如何相互作用。这将使我们能够确定哪些可能的光色素可以赋予通常对光没有反应的细胞光敏性。最后，测量这些光敏蛋白在视网膜或大脑中的真正功能是非常困难的，因为它们只存在于相对较少的细胞中，因此很难找到。因此，我们将使用新技术对斑马鱼进行基因改造，从而使通常产生这些光色素的细胞被改造成荧光标记发光的动物。这些方法将帮助我们探索和理解存在于眼睛和大脑中的所有新的光探测机制。

The Retina is a thin layer of interconnected nerve cells that covers the inside surface of the back of the eyeball. The whole process of Vision itself begins with a small group of proteins (known as opsins) which perform the unique task of absorbing light and translating it into an electrical/biological signal. In the retina, it was traditionally assumed that these opsins were confined to the rod and cone cells, known collectively as photoreceptors. These photoreceptor cells are the functional equivalent of the pixels in the CCD array of a digital camera. It has more recently been discovered that there is actually a rich diversity of additional candidate opsins produced in alternative cells in the retina, however their function remains largely unexplored. Furthermore, these same light sensitive proteins are also found in the brain in places like the pineal organ. Collectively these observations indicate that the whole process of light detection is far more complex than originally thought. Here in the current program we propose a systematic study of these potentially light sensing proteins by looking at the whole diverse opsin photopigment family in the zebrafish, a common model organism. Initially, we will insert the DNA that codes for these proteins into host cells grown in culture conditions. In this way we can measure and compare their functional properties as well as studying how they might interact with one another. This will allow us to define which of these possible photopigments can confer photosensitivity in cells which normally do not respond to light. Finally, it is very hard to measure the real function of these light sensitive proteins in the retina or brain because they are only found in relatively few cells and as a result they are simply very hard to find. We will therefore use new techniques to genetically modify zebrafish, so as to generate animals where the cells that normally produce these photopigments are modified to glow with a fluorescent marker. These approaches will help us explore and understand the whole range of new light detection mechanisms present in the eye and brain.