Mechanism of action of Retinal Determination proteins

描述（由申请人提供）：视网膜血管系统的适当发育对视力至关重要。此外，糖尿病视网膜病变、年龄相关性黄斑变性和早产儿视网膜病变等疾病中血管生长异常是导致失明的常见原因。视网膜决定（RD）级联是一个进化保守的信号通路，在果蝇眼睛发育的背景下得到了最好的研究，包括Eyes Absent （EYA）和Sine oculis homeobox （SIX）家族的蛋白质。我们最近证明，眼缺失在血管发育中起着至关重要的作用。我们现在有证据表明SIX家族转录因子与EYA有机制协同作用，巨噬细胞和内皮细胞中的SIX蛋白都参与调节血管生成。这种竞争性更新应用建立在这些观察的基础上，专门测试SIX蛋白在发育性和病理性视网膜血管生成中的作用。我们假设视网膜髓细胞中的SIX1上调VEGFC的表达，从而促进了邻近视网膜血管的端到柄转换和吻合。内皮细胞中的SIX1/2在小管形成过程中促进分支和增殖，并上调VEGFC的表达，从而加强了尖端到茎的转化。为了在体外和体内验证这一假设，我们提出以下具体目标：(I)阐明巨噬细胞SIX1在调节视网膜血管生成中的作用；（II）阐明内皮细胞SIX1/2在视网膜血管生成中的作用。每个目标都将通过整合各种技术来实现，包括机械生物化学、基于细胞培养的分析、化学生物学和眼睛发育和疾病的动物模型。该项目的实际成果将包括：(1)视网膜血管生成被调节的机制的描述，(2)可以靶向治疗增生性视网膜病变的途径的鉴定和分子表征，以及(3)可以进一步开发为靶向治疗的铅小分子抑制剂的验证。

DESCRIPTION (provided by applicant): Appropriate development of the retinal vasculature is essential for vision. Moreover, abnormal blood vessel growth in diseases such as diabetic retinopathy, age related macular degeneration and retinopathy of prematurity are common causes of blindness. The Retinal Determination (RD) cascade is an evolutionarily conserved signaling pathway best studied in the context of fly eye development and includes proteins of the Eyes Absent (EYA) and Sine oculis homeobox (SIX) families. We have recently demonstrated that Eyes Absent plays an essential role in vascular development. We now have evidence that the SIX family of transcription factors mechanistically synergize with EYA, and that the SIX proteins in both macrophages and in endothelial cells participate in regulating angiogenesis. This competing renewal application builds upon these observations to specifically test the role of the SIX proteins in developmental and pathological retinal angiogenesis. We hypothesize that SIX1 in retinal myeloid cells upregulates VEGFC expression and thus promotes tip-to-stalk conversion and anastomoses in the adjacent retinal vasculature. SIX1/2 in endothelial cells promotes branching and proliferation during tubulogenesis as well as upregulating VEGFC expression, thus reinforcing the tip-to-stalk conversion. To test this hypothesis in vitro and in vivo we propose the following Specific Aims: (I) To elucidate the role of macrophage SIX1 in regulating retinal angiogenesis, and (II) To elucidate the role of endothelial SIX1/2 in retinal angiogenesis. Each aim will be accomplished by integrating diverse techniques including mechanistic biochemistry, cell culture based assays, chemical biology and animal models of eye development and disease. The tangible outcomes of this project will include (1) the delineation of mechanisms by which retinal angiogenesis is regulated, (2) the identification and molecular characterization of a pathway that could be targeted for therapeutic benefit in proliferative retinopathies, and (3) the validation of lead small molecule inhibitors tht could be further developed as targeted therapeutics.