The mechanism of GPCR signalling in zebrafish semicircular canal morphogenesis

内耳的半规管检测我们头部的转动，触发肌肉反射，使我们能够保持平衡。每个内耳都有三个耳道，它们彼此成直角排列，因此它们可以检测任何方向的运动。了解这些管道如何在胚胎中发育，以及内耳的其他复杂结构也被正确地称为外耳道是一个有趣的问题。我们在斑马鱼胚胎中研究了这一过程，但与人类发育过程中的事件序列有许多相似之处。斑马鱼的耳朵在早期胚胎中最初是一个简单的充满液体的细胞球。然后形成组织突起，它们生长到这个球内充满液体的空间中，在那里它们接触并融合形成支柱，成为半规管导管的枢纽。我们最近发现了一种受体蛋白，它似乎参与触发一系列事件，使投射能够相互识别，融合并相应地重新排列。当编码这种受体的基因被破坏时，这些突起就无法接触和融合：它们看起来像是在没有识别的情况下滑过另一个。结果，半规管不能正常形成，耳朵变得畸形和肿胀，鱼长大后有轻微的平衡缺陷。该受体是一类有趣的蛋白质，称为GPCR，位于细胞膜上，从其表面突出。然而，GPCR受体并不是孤立地起作用，而是通常对细胞表面的信号分子做出反应，然后触发细胞内激活新基因的事件。该项目的目的之一是确定这一进程中的其他参与者。我们有一些很好的线索，从鱼类品种，其中半规管未能正确形成。我们的目标是确定参与同一过程的任何新基因。我们还将收集有关受体蛋白何时何地存在，如何调节以及在发育过程中如何发挥作用的信息。我们已经确定了一些受受体调控的基因。其中一类，多能蛋白，在受体功能缺失时仍然高度活跃，而正常情况下它们应该被关闭。我们将测试versicans是否是真正控制突起如何在发育中的斑马鱼耳朵中移动和融合的参与者。最后一个目标是寻找可以影响耳朵半规管形成过程的小药物样分子。有许多GPCR蛋白质，这些蛋白质构成了当今市场上许多药物的靶标。因此，有相当大的商业利益，了解更多关于GPCR和确定新的药物，影响他们的活动。我们的目标是进行一项小规模的研究，以确定新的受体激活剂和抑制剂，然后我们将能够在未来的项目中进一步开发。虽然该项目旨在了解斑马鱼耳朵中发生的过程，但GPCR在胚胎的其他区域也很活跃，因此这些结论可能有助于我们了解其他器官系统的发育，如心脏和神经系统。

The semicircular canals of the inner ear detect turning movements of our head, triggering muscular reflexes that enable us to maintain our balance. There are three canals in each inner ear, and these are each arranged at right angles to one another, so that they can detect movement in any direction. Understanding how these canals develop in the embryo, together with the rest of the intricate structure of the inner ear-rightly also known as the labyrinth-is an interesting problem. We study this process in the zebrafish embryo, but there are many similarities to the sequence of events during human development. The zebrafish ear initially starts out in the early embryo as a simple fluid-filled ball of cells. Projections of tissue then form that grow out into the fluid-filled space inside this ball, where they touch and fuse to make pillars, which become the hubs of the semicircular canal ducts. We have recently identified a receptor protein that appears to be involved in triggering a whole series of events to enable to the projections to recognise one another, fuse and rearrange accordingly. When the gene coding for this receptor is disrupted, the projections fail to touch and fuse: they appear to slide past one another without recognition. As a result, the semicircular canals do not form properly, the ear becomes misshapen and swollen, and the fish grow up with mild balance defects. The receptor is a member of an interesting class of proteins called GPCRs that are located in the cell membrane, projecting from its surface. A GPCR receptor does not act in isolation, however, but usually responds to a signalling molecule at the cell surface, and then triggers events inside the cell that activate new genes. One of the aims of this project is to identify the other players in this process. We have a number of good clues from fish strains in which the semicircular canals fail to form correctly. We will aim to identify any new genes involved in the same process. We will also gather information about when and where the receptor protein is present, how it is regulated, and how it functions during the development process. We have already identified some of the genes that are regulated by the receptor. One class of these, the versicans, remain highly active when receptor function is missing, when they should normally be turned off. We will test whether the versicans are the players that actually control how the projections move and fuse in the developing zebrafish ear. A final aim is to search for small drug-like molecules that can affect the process of semicircular canal formation in the ear. There are many GPCR proteins, and these comprise targets for many of the drugs that are on the market today. There is, therefore, considerable commercial interest in understanding more about GPCRs and in identifying new drugs that affect their activity. We will aim to carry out a small-scale study to identify new activators and inhibitors of the receptor, which we will then be able to develop further in future projects. Although this project aims to understand processes that happen in the zebrafish ear, the GPCR is active in other areas of the embryo, and so the conclusions are likely to aid in our understanding of the development of other organ systems, such as the heart and nervous system.