Elucidating the role of DNA methyltransferases in epigenetic regulation of retinal regeneration in zebrafish

项目摘要 根据国家眼科研究所的数据，目前有超过1200万美国人患有 来自影响视网膜的疾病。提出，治疗尝试减少视网膜死亡或 替换丢失的神经元取得的成功有限，这突显了寻找替代方案的必要性 解决这个问题的方法。斑马鱼正在成为一个越来越受欢迎的研究模型 干细胞为基础的组织再生机制。为了应对广泛的视网膜损伤， 斑马鱼能够完全再生视网膜。这是通过归纳法实现的 进行不对称分裂以产生祖细胞池的Müler神经胶质细胞 继续再生所有类型的视网膜细胞，没有观察到胶质瘢痕的证据 在哺乳动物物种中。而斑马鱼再生组织的非凡能力 多年来，我们对哺乳动物和哺乳动物之间的比较生物学的理解 硬骨鱼对视网膜损伤的反应仍然知之甚少。目前，视网膜的研究 斑马鱼的再生主要集中在识别信号通路和 参与视网膜再生的单个基因。这些途径的表观遗传编排， 然而，斑马鱼中的这一现象仍有待研究。DNA甲基转移酶(DNMT)已被 在哺乳动物中进行了广泛的研究，并已被确定为动态平衡的关键 维持成体干细胞进程，如造血干细胞发育。这个 有关斑马鱼表观遗传调节因子的有限文献显示，硬骨鱼物种似乎利用 斑马鱼Dnmts与哺乳动物物种相同的表观遗传调控机制 与哺乳动物Dnmts有相当大的序列同源性。据了解，即刻 斑马鱼视网膜损伤后，初始的全局DNA低甲基化和 随后，随着祖细胞池开始分化，DNA甲基化增加， 这表明了表观遗传景观在这些过程中的重要性。与 在这项提案中计划的实验，我们的目的是确定DNMT在协调 斑马鱼成体视网膜再生过程，目的在于阐明再生途径 在表观遗传水平上调控在Müler神经胶质来源的祖细胞中。我们还计划开发 一个新的工具，将是斑马鱼群落的重要补充，用于调查 特定基因的有针对性的表观遗传调控的后果。这些拟议的研究将 阐明靶向治疗方法的候选基因以识别和“解锁”通路 在表观遗传沉默的哺乳动物系统中，为未来的研究开辟了新的途径 在再生医学领域。

Project Abstract According to the National Eye Institute, there are currently over 12 million Americans suffering from diseases affecting the retina. To present, therapeutic attempts to reduce retinal death or replace lost neurons have been met with limited success, highlighting the need for an alternative approach to this problem. The zebrafish is becoming an increasingly popular model to study mechanisms of stem-cell based tissue regeneration. In response to extensive retinal injury, zebrafish are able to completely regenerate the retina. This is accomplished through the induction of Müller glial cells which undergo an asymmetric division to generate a pool of progenitor cells that go on to regenerate all cell types of the retina with no evidence of the glial scarring observed in mammalian species. While the remarkable capacity of the zebrafish to regenerate tissues has been known for years, our understanding of the comparative biology between mammalian and teleost responses to retinal damage remains poorly understood. To present, studies of retinal regeneration in the zebrafish have largely focused on identifying signaling pathways and individual genes involved in retinal regeneration. The epigenetic orchestration of these pathways, however, remains to be investigated in the zebrafish. DNA methyltransferases (Dnmts) have been studied extensively in mammals and have been identified as critical in the homeostatic maintenance of adult stem cell processes such as hematopoietic stem cell development. The limited literature of epigenetic regulators in zebrafish reveals that teleost species appear to utilize the same mechanisms of epigenetic regulation as mammalian species, with zebrafish Dnmts baring considerable sequence homology to mammalian Dnmts. It is known that immediately following retinal injury in the zebrafish there is an initial global DNA hypomethylation and subsequent increase in DNA methylation as the pools of progenitor cells begin to differentiate, indicating the importance of the epigenetic landscape during these processes. With the experiments planned in this proposal, we aim to identify the role of Dnmts in orchestrating the process of adult retinal regeneration in the zebrafish, with the goal of elucidating pathways regulated in Müller glial derived progenitor cells at the epigenetic level. We also plan to develop a new tool that will be an important addition to the zebrafish community for investigation of the consequences of targeted epigenetic modulation of specific genes. These proposed studies will illuminate candidate genes for targeted therapeutic approaches to identify and “unlock” pathways in mammalian systems that are epigenetically silenced, opening new avenues for future studies in the field of regenerative medicine.