"Role of Pitx3 signaling cascades in the regulation of Xenopus eye, laterality, and somite development"

令人惊讶的是，我们对人类眼睛发育的了解大多是在对两栖动物胚胎的研究和操作中得到的。发育、结构和基因的相似性是惊人的：不管你是研究人类、老鼠、鱼还是青蛙，相同的基因相互作用产生相似的结构。例如，人类PITX3同源盒基因突变导致晶状体不能形成或先天性白内障。同样的问题可以在发育中的老鼠、青蛙甚至鱼的胚胎中重现，其中涉及的基因和过程可以被研究和操纵。研究青蛙基因的好处是，胚胎相对较大，它们在外部发育（你可以看到它们的地方），而且它们的操作成本很低。非洲爪蟾的基因组已经接近测序，并且有许多有用的探针和技术来研究该物种的眼睛发育，因此该生物对于检验我们关于人类眼睛发育和疾病的理论是有用的。在我们最近对Pitx3在青蛙中的作用的研究中，我们发现该基因不仅在眼睛发育中起作用，而且在器官不对称（心脏、胆囊、肠道）和椎骨中也起作用。我们已经确定了Pitx3控制的一些基因，这些基因在建立胚胎体计划中发挥作用，并调节一些对细胞形状、结构和活动重要的过程。我们已经制定了策略来评估Pitx3信号级联如何展开以控制眼睛的发育和健康，以及内脏器官和脊柱的发育议程。我们正在申请资金来继续这项工作，并研究Pitx3如何开启一些下游目标基因，同时关闭其他基因。我们希望更好地理解一个基因的活动如何安排一个发展议程。我们使用基于流式细胞术的方法来评估Pitx3对单个细胞和基因的影响的初步工作已经产生了许多下游信号级联，我们需要测试和操纵这些信号级联，以完善我们对脊椎动物胚胎中早期事件如何调节的理解。

Surprisingly, much of what we have learned about human eye development was first pioneered in studies and manipulations of amphibian embryos. The developmental, structural, and genetic similarities are striking: irrespective of whether you study human, mouse, fish or frog, the same genes interact to produce similar structures. For example, mutation of the human PITX3 homeobox gene results in a failure of lenses to form or in congenital cataracts. The same issues can be re-capitulated in developing mouse, frog, and even fish embryos, where the genes and processes that are involved can be studied and manipulated. The benefit of studying the genes in frog is that the embryos are comparatively large, they develop externally (where you can see them), and they can be cheaply manipulated. The genome is nearly sequenced for the frog Xenopus, and there are many useful probes and techniques for studying eye development in this species, so the organism is useful for testing our theories regarding human eye development and disease. During our recent studies of the role of Pitx3 in frog, we discovered that the gene plays a role not only in eye development, but also in organ asymmetry (heart, gall bladder, gut), and vertebrae. We have identified some of the genes that Pitx3 controls, and these genes play a role in establishing the embryonic body plan, as well as regulating some of the processes important to cell shape, structure, and activity. We have developed strategies to assess how the Pitx3 signaling cascade unfolds to control development and health of the eye, as well as the agenda for development of visceral organs and spine. We are requesting funds to continue this work and to study how Pitx3 turns on some downstream target genes, while turning off others. We hope to reach a better understanding of how the activity of one gene installs an agenda for development. Preliminary work using our newly developed protocol using a flow cytometry based approach to asses Pitx3 effects upon individual cells and genes has produced a number of downstream signaling cascades that we need to test and manipulate in order to refine our understanding of how early events are regulated in vertebrate embryos.