A newly discovered feed-forward mechanism controls photoreceptor fate

描述(申请人提供)：视网膜变性是一种破坏性的疾病，由于视网膜最外层的视杆细胞和视锥感光细胞的进行性死亡而导致不可逆转的视力丧失。在退化后更换光感受器的最有希望的途径之一是将光感受器前体细胞引入视网膜，这些细胞可以分化为视杆和视锥。然而，在对前体的试验中， 移植治疗时，会产生太多的杆状细胞，而不是足够的锥体。这些试验的结果揭示了使用干细胞疗法恢复视力的一个主要障碍，并突显了我们在光感受器形成方面的知识中的一个关键缺口：我们不知道将前体从首先产生的锥状光感受器转换到产生棒状光感受器的机制。为了了解视锥和视杆感光器的数量是如何调节的，我们建立了一个简单但强大的工具来研究光感受器的发育，斑马鱼松果体与眼睛有着非常密切的进化关系。就像在视网膜中一样，斑马鱼的松果体含有完全发育的视杆。 此外，松果体的光感受器在分子上与视网膜中的光感受器非常相似。斑马鱼松果体研究光感受器发育的一个主要优势是它的简单，可以在整个胚胎中轻松地对活的或固定的松果体器官进行成像，而不需要切片，因为没有晶状体、房水或覆盖的视网膜层的干扰。我们实验室最近的研究表明了一种新的和意想不到的机制，称为非相干前馈系统，它控制光感受器前体产生的视锥细胞和视杆细胞的数量。我们认为，在产生视锥细胞的早期光感受器前体中，转录因子Tbx2b的限时表达驱动了视锥特异基因的表达，也启动了视锥基因抑制因子Nr2e3的表达。Nr2e3蛋白继续积累，直到它达到一个阈值浓度，关闭CONE基因的表达，并允许杆状结构的形成。为了测试我们的前馈假说对光感受器形成的影响，我们将利用我们和其他人在斑马鱼中产生的遗传工具。然后，我们将利用我们的知识来测试Nr2e3致病突变对前馈控制的影响。我们的研究将阐明视锥和视杆产生的分子控制，这将为实验室开发视网膜替代疗法以及深入了解遗传性视网膜变性疾病的个体发育提供宝贵的信息。

DESCRIPTION (provided by applicant): Retinal degeneration is a devastating affliction that leads to irreversible loss of vision, resulting from the progressive death of rod and cone photoreceptor cells in the outermost layer of the retina. One of the most promising avenues for replacing photoreceptors following degeneration is the introduction of photoreceptor precursor cells into the retina, which can differentiate into rods and cones. However, in trials of precursor transplant therapy, too many rods and not enough cones are produced. The result of these trials reveals a major hurdle for restoration of vision using stem-cell therapies, and highlights a criticl gap in our knowledge of photoreceptor formation: We do not know the mechanism that switches precursors from production of cone photoreceptors, which arise first, to production rod photoreceptors, which arise second. In order to understand how the number of cone versus rod photoreceptors is regulated, we have established a simple but powerful tool to study photoreceptor development, the zebrafish pineal organ, which shares a very close evolutionary relationship with the eyes. Just as in the retina, the zebrafish pineal contain fully developed rod and cone photoreceptors; in addition, the pineal photoreceptors are molecularly quite similar to those in the retina. A major advantage of the zebrafish pineal for studying photoreceptor development is its simplicity, which allows imaging of the live or fixed pineal organ easily in whole mount embryos without sectioning since there is no interference from the lens, aqueous humor, or overlying retinal layers. Recent studies from our laboratory indicate a novel and unexpected mechanism, called an incoherent feed-forward system, which controls the number of cone and rod cells produced by photoreceptor precursors. We propose that in early photoreceptor precursors, which give rise to cone cells, time-limited expression of the transcription factor Tbx2b expression drives the expression of cone-specific genes and also initiates expression of the cone-gene repressor Nr2e3. Nr2e3 protein continues to accumulate until it reaches a threshold concentration that shuts off cone gene expression and allows rods to form instead. In order to test the implications of our feed-forward hypothesis for photoreceptor formation, we will take advantage of the genetic tools that we and others have generated in zebrafish. Then we will leverage our knowledge to test the effects of a disease-causing mutation in Nr2e3 on feed-forward control. Our studies will illuminate the molecular control of cone versus rod production, which will provide invaluable information for laboratories developing retinal replacement therapies as well as insight into the ontogeny of inherited retinal degeneration disorders.