The mechanism of GPCR signalling in zebrafish semicircular canal morphogenesis

内耳的半圆形通道探测到我们头部的转动，触发肌肉反射，使我们能够保持平衡。每只内耳都有三条通道，它们彼此成直角排列，因此它们可以探测到任何方向的运动。了解这些管道是如何在胚胎中发育的，以及内耳的其他复杂结构——也被正确地称为迷宫——是一个有趣的问题。我们在斑马鱼胚胎中研究了这一过程，但在人类发育过程中有许多相似之处。斑马鱼的耳朵最初是在胚胎早期开始的，是一个简单的充满液体的细胞球。然后形成组织的突起，生长到球内充满液体的空间，在那里它们接触并融合成柱子，成为半圆形管道的枢纽。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，使突起能够相互识别，融合并相应地重新排列。当这种受体的基因编码被破坏时，投射物就无法接触和融合：它们似乎彼此滑过而不被识别。因此，半规管不能正常形成，耳朵变得畸形和肿胀，鱼长大后会有轻微的平衡缺陷。这种受体是一类有趣的蛋白质的一员，这种蛋白质被称为gpcr，位于细胞膜上，从细胞膜表面伸出。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作出反应，然后在细胞内触发激活新基因的事件。这个项目的目的之一是确定这个过程中的其他参与者。我们从鱼种中得到了一些很好的线索，其中半规管不能正确形成。我们的目标是找出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。当受体功能缺失的时候，当它们通常应该被关闭的时候，其中一类，versicans，仍然保持高度活跃。我们将测试这些变体是否是真正控制投影如何移动和融合在发育中的斑马鱼耳朵中的玩家。最后的目标是寻找能影响耳朵中半规管形成过程的类似药物的小分子。有许多GPCR蛋白，它们构成了当今市场上许多药物的靶标。因此，更多地了解GPCRs和确定影响其活性的新药具有相当大的商业利益。我们的目标是开展一项小规模研究，以确定受体的新激活剂和抑制剂，然后我们将能够在未来的项目中进一步开发。虽然这个项目旨在了解斑马鱼耳朵中发生的过程，但GPCR在胚胎的其他部位也很活跃，因此结论可能有助于我们理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.