Identification of factors controlling the genesis of cone photoreceptors

描述(申请人提供)：这里描述的研究旨在阐明视网膜中一种特定类型的神经元-视锥感光细胞产生的分子机制。控制这种特殊细胞类型发展的因素还不是很清楚。通过对一种产生过量锥体的小鼠突变体的微阵列分析，我们的实验室已经确定了一些与这种细胞效应有关的上调基因。这些基因中有两个是锥体转导蛋白亚基：Gnat2，G蛋白α亚基，和Gnb3，G蛋白β亚基。在成年脊椎动物视网膜中，GNat2仅在视锥感光细胞中表达，而Gnb3则在双极细胞和视锥细胞中表达。在这里，它们的功能是在视蛋白下游的信号转导级联中传递光刺激，视蛋白是G蛋白偶联的受体。本研究提出的初步证据表明，GNat2和Gnb3在小鼠和鸡的早期光感受器层中都有特异性表达，这是通过RNA原位杂交检测的。与其他转导途径成分相比，这种表达是早熟的，大多数转导途径成分直到发育很晚才表达。这项提议将测试这些G蛋白亚基是否在发育早期发挥功能，以确定锥体的命运，或者允许锥体光感受器的适当分化。功能丧失和功能获得的研究将在鸡的视网膜上进行，以测试这些基因在视锥细胞生产中的必要性和/或充分性。一个候选的G蛋白偶联受体家族可以在发育早期发挥Gnat2和Gnb3上游的功能，这就是Frizzled型受体家族。Gnat2已被证明与Frizzled2受体偶联，以响应Frizzled2受体的分泌配体WntSa。实验将被设计来扩展之前的发现，即Wnt信号参与了锥体的发育。最终的目标将是转录描述早期视锥细胞光感受器谱系，以确定其他候选基因，这些基因具有产生完全分化的视锥细胞的功能。我们期待这些研究为人类视觉中使用的主要光感受器--视锥细胞--如何在发育过程中获得新的见解提供新的见解。将要测试的候选光感受器发生基因之一(Gnat2)已被确定为色视(无色觉)的遗传原因，这项提议将解决这种疾病的机制。至少有2000万至2500万人受到遗传病的困扰，这些疾病会导致光感受器退化和视力下降。对产生光感受器的正常机制的基本了解将为用新的功能光感受器取代这些细胞提供基础。

DESCRIPTION (provided by applicant): The studies described here are designed to elucidate the molecular mechanisms underlying the production of a specific class of neurons - cone photoreceptors of the retina. The factors controlling the development of this specialized cell type are not well understood. Using microarray analysis of a mouse mutant that produces excess cones, our lab has identified a number of genes upregulated concommitantly with this cellular effect. Two of these genes are the cone transducin subunits: Gnat2, a G-protein alpha subunit, and Gnb3, a G-protein beta subunit. In the adult vertebrate retina, Gnat2 is expressed exclusively in cone photoreceptors, while Gnb3 is found in bipolar cells and cones. Here they function to transduce light stimulus in a signal transduction cascade downstream of opsins, which are G-protein coupled receptors. Preliminary evidence presented in this proposal shows that Gnat2 and Gnb3 are both expressed specifically in the early photoreceptor layer in both mice and chickens as assayed by RNA in situ hybridizations. This expression is precocious as compared to the other transduction pathway components, most of which are not expressed until much later in development. This proposal will test whether these G-protein subunits function early in development to specify the cone fate or allow for the proper differentiation of cone photoreceptors. Loss-of-function and gain-of-function studies will be used in the chicken retina to test the necessity and/or sufficiency of these genes in the production of cones. A candidate G-protein coupled receptor family that could function upstream of Gnat2 and Gnb3 early in development is the Frizzled family of receptors. Gnat2 has been shown to couple to Frizzled2 receptor in response to WntSa, a secreted ligand for Frizzled receptors. Experiments will be designed to extend previous findings that Wnt signaling is involved in cone development. The final aim will be to transcriptionally profile the early cone photoreceptor lineage to identify other candidate genes that function to generate fully differentiated cones. We envision these studies providing new insights into how the primary photoreceptor used in human vision, the cone, is derived during development. One of the candidate photoreceptor-genesis genes that will be tested (Gnat2) has been identified as a genetic cause of achromatopsia (no color vision) and this proposal will address the mechanism for this disease. At least 20 to 25 million people are afflicted with genetic diseases that cause the degeneration of photoreceptors and a decrease in their vision. A basic understanding of the normal mechanisms that generate photoreceptors will provide the basis for replacing these cells with new functional photoreceptors.