Transcriptional Regulation of Cone Photoreceptor Genesis

描述(申请人提供)：视锥感光细胞(视锥细胞)在人类视觉中起着至关重要的作用，如果这些细胞死亡，就会出现严重的视力障碍。例如，视网膜色素变性和黄斑变性是两种这样的疾病，在这些疾病中，视力的严重丧失与视锥细胞的丧失有关。治疗这些疾病的一种有希望的疗法是用患者体内或外部产生的新锥体取代丢失的锥体。然而，脊椎动物圆锥体发生的基本分子机制仍然不清楚。如果没有这些信息，生产圆锥体的合理策略是不可用的。长期目标是确定促进锥体发生的基因调控网络，并在此基础上设计方法来制造新的锥体细胞用于细胞替代治疗。这项提案的总体目标是从功能上剖析转录网络 允许特定的视网膜祖细胞亚群优先产生两种罕见的视网膜细胞类型，视锥细胞和水平细胞。这一提议将检验核心假设，即 转录因子OTX2和ONECUT1是这种特殊的锥体/水平细胞前体细胞的主要调节因子。开展这项研究的基本原理是，通过了解促进这些祖细胞和最终锥体感光细胞形成的转录因子网络，在体外或体内产生新的锥体感光细胞的方法将成为可能。为了验证这一假设，提出了两个特定的目标：1)确定建立视锥/水平祖细胞状态和促进视锥发生的转录因子；2)确定诱导视锥/水平祖细胞和从其他类型的视网膜祖细胞分化的视锥细胞的关键参数Onecut1和OTX2的表达。第一个目标将确定在锥体/水平祖细胞中特异表达的基因，以及这种分子签名在多大程度上是由Onecut1和OTX2转录因子产生的。此外，它还将确定抑制杆状感光细胞发生的转录因子途径。第二个目标是测试Onecut1和OTX2在多大程度上可以重新编程其他视网膜前体细胞，使其产生视锥细胞和水平细胞，并最终驱动视锥分化程序，同时抑制视杆细胞的形成。这种方法是创新的，因为它将在整个转录组水平上从功能上研究视网膜祖细胞的特定亚群。此外，t还将剖析和探索参与锥体发生的最上游转录因子网络。在这个项目完成后，将有新的途径来了解特定视网膜前体细胞类型的生物学，以及视锥和视杆感光细胞发育的早期步骤。这里提出的这项研究意义重大，因为它有望揭示视锥体光感受器起源的新的、基本的见解。预计这一知识将对开发治疗人类失明的新疗法具有翻译潜力。

DESCRIPTION (provided by applicant): Cone photoreceptors (cones) serve a critical function in human vision that is underscored by the dramatic impairments in sight that occur if these cells die. For instance, retinitis pigmentosa and macular degeneration are two such diseases in which substantial loss of vision correlates with the loss of cones. One promising therapy for these diseases is to replace lost cones with new ones produced inside or outside of the patient. However, the fundamental molecular mechanisms underlying the genesis of vertebrate cones are still unclear. Without this information, a rational strategy to produce cones is unavailable. The long-term goal is to identify the gene regulatory networks that promote cone genesis, and to devise methods based on this knowledge to make new cone cells for cell replacement therapy. The overall objective of this proposal is to functionally dissect the transcriptional networks that allow a specific sub-population of retinal progenitor cells to preferentially generate two rare retinal cell types, cones and horizontal cells. This proposal will test the central hypothesis that the transcription factors Otx2 and Onecut1 are the master regulators of this specific cone/horizontal cell progenitor cell. The rationale for undertaking this study is that by understanding the transcription factor networks that promote the formation of these progenitor cells and ultimately cone photoreceptor cells, methods for generating new cone photoreceptors in vitro or in vivo will become possible. To test this hypothesis, two specific aims are proposed: 1) Identify the transcription factors that establish the cone/horizontal progenitor cell state and promote cone genesis and 2) Define critical parameters of Onecut1 and Otx2 expression that induce cone/horizontal progenitors and differentiated cones from other retinal progenitor cell types. The first aim will determine the genes expressed specifically in cone/horizontal progenitor cells and the extent to which this molecular signature is generated by the Onecut1 and Otx2 transcription factors. In addition, it will identify the transcription factor pathways downstream o Onecut1 that repress rod photoreceptor genesis. The second aim will test the extent to which Onecut1 and Otx2 can reprogram other retinal progenitors into the specific type that generates cones and horizontal cells and ultimately drive the cone differentiation program, while repressing rod genesis. This approach is innovative because it will functionally investigate a specific sub-population of retinal progenitor cells at the whole transcriptome level. In addition, t will dissect and explore the most upstream transcription factor network involved in cone genesis. At the completion of this project, new avenues will be available to understand the biology of specific retinal progenitor types, as well as the early steps in cone and rod photoreceptor development. The research proposed here is significant because it is expected to reveal novel, fundamental insights into the genesis of cone photoreceptors. It is expected that this knowledge will have translational potential for the development of new therapies for human blindness.