A role for a neprilysin in neuronal differentiation during retinal development

描述(由申请人提供)：了解增殖和分化之间的转变对许多领域具有重要意义，包括基于干细胞的视网膜变性治疗。在果蝇和脊椎动物的视网膜发育过程中，从增殖前体细胞到分化神经元的转变是在一个波中发生的，该波通过视网膜前体组织传播，并由Hedgehog(HH)和其他信号通路之间的相互连接控制。波的传播与神经发育所必需的无音(ATO)相关神经因子的表达有关。我们已经确定了一种新的基因，异常水泡和畸形眼(ABAM)，它需要协调波的传播并将其与果蝇眼的神经分化联系起来。令人惊讶的是，在缺乏R8的情况下，当ABAMS功能降低时，神经分化就会发生，尽管是以一种混乱的方式。这种表型不同于以前观察到的任何一种表型，表明我们在控制波传播的信号通路是如何协调并与神经分化相关的知识方面存在显著差距。Abams基因编码II型跨膜金属内肽酶neprilysin家族的一个预测成员。在脊椎动物中，neprilysins裂解信号肽，例如调节血压，并在年龄相关性黄斑变性的背景下与Aβ代谢有关，但对它们在调节发育信号中的功能知之甚少。有趣的是，阿巴姆斯缺乏金属肽酶催化活性的关键残基。一些哺乳动物的Neprilysins(如苯丙氨酸)结合到底物上，并防止其他多肽酶的降解。根据我们的初步证据，我们假设ABAMS结合并阻止信号因子的切割，该信号因子协调波的传播，并将其与视网膜发育过程中的神经分化联系起来。这项工作的长期全球目标是了解在视网膜发育背景下发生增殖和分化之间转换的机制。本申请的目的是确定哪些信令事实或因素是ABAMS的目标。为了实现这些目标，我们将(1)确定ABAMS在哪些细胞中起作用以及它在细胞中的定位；(2)确定ABAMS对波传播重要的信号通路的影响；(3)确定ABAMS与哪些蛋白质相互作用。这里提出的实验有望导致识别ABAMS靶向的信号通路成分，以及neprilysins如何在眼睛中发挥作用。如果成功，这项工作将使未来能够进行实验，了解信号通路如何控制神经元分化的本质，以及abams和潜在的其他neprilysins在眼睛发育和功能中的作用。

DESCRIPTION (provided by applicant): Understanding the transition between proliferation and differentiation has important implications for many fields, including for stem cell-based therapies for retinal degeneration. During retinal development in both Drosophila and vertebrates the transition from proliferating precursors to differentiating neurons occurs in a wave that propagates across retinal precursor tissue and is controlled by interconnections between Hedgehog (Hh) and other signaling pathways. Wave propagation is linked to expression of Atonal (Ato)-related proneural factors, which are necessary for neural development. We have identified a novel gene, abnormally blistered and misshapen eyes (abams), that is required to coordinate wave propagation and link it to neural differentiation in the Drosophila eye. Surprisingly, neural differentiation occurs in the absence of R8 when abams function is reduced, albeit in a disorganized fashion. This phenotype is unlike any that has been observed previously, and suggests a significant gap in our knowledge of how the signaling pathways that control wave propagation are coordinated and linked to neural differentiation. The abams gene encodes a predicted member of the neprilysin family of type II transmembrane metalloendopeptidases. In vertebrates neprilysins cleave signaling peptides that e.g. regulate blood pressure and have been linked to Aβ metabolism in the context of age related macular degeneration, but little is known about their functions in regulating developmental signals. Interestingly, Abams lacks residues critical for metallopeptidase catalytic acitivity. Some mammalian neprilysins (e.g. PHEX) bind to substrates and prevent degradation by other peptidases. Based on our preliminary evidence, we hypothesize that Abams binds to and prevents cleavage of a signaling factor that coordinates wave propagation and links it to neural differentiation during retinal development. The long-term global aim of this work is to understand the mechanisms by which the transition between proliferation and differentiation occurs in the context of retinal development. The object of this application is to identify which signaling facto or factors is targeted by Abams. To accomplish these goals we will (1) determine in which cells Abams acts as well as its localization in cells; (2) determine the effect of Abams on signaling pathways important for wave propagation and (3) Determine what proteins Abams interacts with. The experiments proposed here are expected to lead to identification of the signaling pathway component targeted by Abams and how neprilysins function in the eye. If successful, this work will enable future experiments into the nature of how signaling pathways control neuronal differentiation, and into the role of Abams and potentially other neprilysins in eye development and function.