Essential functions of PITX2 in cornea, iris, and iridocorneal angle development

描述(申请人提供)：我们的目标是确定控制眼前段发育的遗传网络，并了解这些网络的破坏是如何导致眼前节结构和功能缺陷的。我们的目标是测试同源结构域转录因子PITX2在前段组织中的功能，这些组织在人类PITX2突变患者中最常见的影响。我们的中心假设是，PITX2调节正常角膜细胞命运所需的遗传网络，排除发育和成熟的角膜的血管，并调节虹膜和虹膜角膜角结构内的细胞增殖和谱系特征。我们的假设是基于对临时基因敲除小鼠进行分析而产生的初步数据。这项研究的理论基础是，了解PITX2在角膜、虹膜和虹膜角膜角度正常发育过程中调控的遗传网络将促进我们对眼前段发育不良及其相关病理的理解，包括眼压升高。我们将通过追求三个具体目标来验证我们的假设：1)测试关于Pitx2是规范和维持角膜细胞命运所必需的假设，2)测试关于Pitx2对于防止血管生长进入发育和成熟的角膜的假设，以及3)测试对于虹膜和虹膜角膜角的结构的正常发育所必需的Pitx2的假设。根据前两个目标，申请人手中已经可行的时间敲除策略将被用于在角膜发育开始时消融PITX2，之后将使用成熟的方法评估角膜谱系和血管生长。根据第三个目标，在虹膜和虹膜角膜角发育开始时，将使用类似的时间敲除方法，在申请人手中也被确定为可行的方法，以消融Pitx2，并将确定对结构发育的后果。预期的结果是，PITX2在角膜、虹膜和虹膜角膜角度发育中的基本功能将被确定。这种方法是创新的，因为它利用时间基因敲除策略来克服全局和组织特异性的Pitx2基因敲除动物的限制，从而使我们能够研究重要的眼前段后来形成的结构。这项拟议的研究意义重大，因为它将在垂直方向上推进和扩大对眼前段结构在发育过程中如何形成的理解。最终，这些知识将为眼前段发育不全如何导致视力丧失提供深入的见解。

DESCRIPTION (provided by applicant): Our goal is to identify the genetic networks governing ocular anterior segment development and to understand how disruption of these networks leads to defects in anterior segment structure and function. Our objective is to test the functions of the homeodomain transcription factor PITX2 in anterior segment tissues that are most commonly affected in human patients with PITX2 mutations. Our central hypothesis is that PITX2 regulates genetic networks that are required to specify normal corneal cell fates, exclude blood vessels from the developing and mature cornea, and regulate cell proliferation and lineage specification within the iris and structures of the iridocorneal angle. Our hypothesis was formulated on the basis of preliminary data generated by analysis of temporal knockout mice. The rationale for the proposed research is that knowledge of the genetic networks regulated by PITX2 in normal development of the cornea, iris, and iridocorneal angle will advance our understanding of anterior segment dysgenesis and associated pathologies, including elevated intraocular pressure. We will test our hypothesis by pursuing three specific aims: 1) Test the hypothesis that Pitx2 is required for specification and maintenance of corneal cell fates, 2) Test the hypothesis that Pitx2 is required to prevent vascular growth into the developing and mature cornea, and 3) Test the hypothesis that Pitx2 is required for normal development of the iris and structures of the iridocorneal angle. Under the first two aims, a temporal knockout strategy, which is already feasible in the applicant's hands, will be used to ablate Pitx2 at the beginning of corneal development, after which corneal lineages and vascular growth will be assessed using well-established approaches. Under the third aim, an analogous temporal knockout approach, also established as feasible in the applicant's hands, will be used to ablate Pitx2 at the beginning of iris and iridocorneal angle development and the consequences on development of the structures will be determined. The expected outcome is that essential functions of PITX2 in the development of the cornea, iris, and iridocorneal angle will be identified. The approach is innovative because it utilizes a temporal gene knockout strategy to overcome the limitations of global and tissue-specific Pitx2 knockout animals, thereby permitting us to study important later-forming structures in the anterior segment. The proposed research is significant because it will vertically advance and expand understanding of how anterior segment structures are formed during development. Ultimately, such knowledge will provide insights into how anterior segment dysgenesis contributes to vision loss.