Membrane contact sites between endolysosomes and the ER as novel hubs in Ca2+ signalling.

将我们的细胞与细菌细胞区分开来的一个关键特征是细胞器的存在-致力于执行特定任务的微小膜结合结构。但这种分工是有代价的，即细胞器之间需要相互沟通，才能使细胞作为一个整体正常运作。一种新兴的细胞器“颤振”的方式是通过膜接触点。这些位点是细胞器通过桥连接在一起的区域，以便交换小分子，如脂质和钙。了解这些位点的组成以及它们是如何被调节的，对于理解细胞功能是如何协调的至关重要。在这个应用中，我们关注的是内质网（ER）和内溶酶体之间的接触位点，内质网是制造新细胞物质的地方，内溶酶体是吸收、回收和降解物质的地方。值得注意的是，这两种细胞器也是钙的储存库。钙作为一种对骨骼和牙齿很重要的矿物离子，可能是大家最熟悉的。这当然是真的。然而，人们不太熟悉的是它在信号传递中的关键作用。当细胞受到刺激时，细胞内的钙含量迅速增加，这种增加引发了一系列事件，这些事件在很大程度上改变了细胞的行为。这可能对从受精到心脏跳动的所有细胞事件都至关重要。在许多情况下，这些钙来自商店。现在清楚的是，钙从内溶酶体释放“触发”进一步从内质网释放钙——细胞器协同工作的一个很好的例子。不太清楚的是，这是如何发生的，这让我们得出了我们的假设，即这发生在两个细胞器之间的接触点。为此，我们现在已经成功地定义了一些参与连接内溶酶体和内质网的蛋白质，并表明破坏这些接触会损害两个细胞器之间的钙信号。令人惊讶的是，我们还发现接触是由钙本身调节的。我们将在这些发现的基础上，结合研究钙（Patel）和接触点（Futter）的申请人的专业知识。首先，我们将研究内溶酶体上的一种蛋白质，我们认为这种蛋白质会引导钙来稳定接触。我们还将干扰接触并研究当钙从内溶酶体释放以不同方式被激活时，这如何影响钙的变化。在课程的第二部分，我们将研究我们认为是额外接触位点成分的新蛋白质。最后，我们将研究破坏接触对细胞行为的影响。具体来说，我们将研究接触如何影响受体蛋白的功能，受体蛋白参与接收来自细胞外的信息。由于接触点面积小，研究它们并不容易。因此，在这个项目中，我们将努力开发新的方法来促进学习。这将与科学界的许多人有关。该项目的成功结果将为我们提供关于细胞如何控制和使用钙的关键新见解。钙的紊乱已经与许多疾病有关，包括帕金森病和癌症。有缺陷的接触点可能是一个未被发现的罪魁祸首吗？了解钙如何调节膜接触部位，反之亦然，可以为制定药物来建立或破坏接触提供基础。

One critical feature which distinguishes our cells from those of bacteria is the presence of organelles - tiny membrane bound structures dedicated to performing specific tasks. But this division of labour comes at a price namely the need for the organelles to communicate in order for the cell to function properly as a whole. One emerging means by which organelles "chatter" is through membrane contact sites. These sites are regions where the organelles come together via bridges so as to allow exchange of small molecules such as lipids and calcium. Understanding the composition of these sites and how they are regulated is critical for understanding how cell function is coordinated.In this application, we focus on contact sites between the endoplasmic reticulum (ER) where new cellular material is made and endolysosomes that take up, recycle and degrade material. Notably, both of these organelles are also stores of calcium. Calcium is perhaps most familiar as a mineral ion important for bones and teeth. This is certainly true. However, less familiar is its critical role in signalling. When cells are stimulated, calcium levels within the cell rapidly increase and this increase sets in train a series of events which change the behaviour of the cell in significant ways. This is critical for probably all cellular events from fertilisation through to the beating of the heart. In many cases this calcium comes from stores.What is now clear is that release of calcium from endolysosomes "triggers" further calcium release from the ER - an excellent example of organelles working together. What is not so clear is how this occurs leading us to our hypothesis that this happens at contact sites between the two organelles. To this end, we have now succeeded in defining some of the proteins involved in connecting endolysosomes and the ER and shown that disrupting these contacts impairs calcium signalling between the two organelles. Surprisingly, we also find that contacts are regulated by calcium itself.We will build on these findings by combining the expertise of the applicants in studying calcium (Patel) and contact sites (Futter). First, we will study a protein on the endolysosomes which we think channels the calcium to stabilize contacts. We will also interfere with the contacts and study how this impacts calcium changes when calcium release from endolysosomes is activated in different ways. In the second part of the program, we will study new proteins that we think are additional contact site components. Finally, we will examine the consequences of disrupting contacts on cell behaviour. Specifically, we will study how contacts affect the function of receptor proteins that are involved in receiving messages from outside of the cells. It is not easy to study contact sites because of their small size. So within the program we will strive to develop new methods to facilitate study. This will be of relevance to many in the scientific community.The successful outcome of the project will provide us with key new insight into how cells control and use their calcium. Disturbances in calcium have already been linked to many disorders including Parkinson's disease and cancer. Could defective contact sites be an unrecognized culprit? Understanding how calcium regulates membrane contacts sites and vice versa could provide the basis for formulating drugs to make or break contact.