14-3-3 Dependent targeting of K2P channels: A fundamental pathway for the intracellular vesicular trafficking of cell surface membrane proteins

细胞通过其表面存在的蛋白质(受体、离子通道和转运体)对周围环境做出反应并与其进行交流。受体嵌入细胞膜，使细胞能够识别并响应导致细胞反应的特定信号。离子通道和转运体还通过控制离子流入和流出细胞，在控制细胞对刺激的反应方面发挥作用。这些膜蛋白在细胞表面的数量和位置会极大地影响细胞对外界信号的反应，而这些关键的细胞表面蛋白的错误定位可能会导致糖尿病和囊性纤维化等疾病。该项目将研究一种细胞控制细胞表面膜蛋白数量的机制。细胞有多种方法来监测和控制其表面膜蛋白的数量和类型。蛋白质携带信号基序(类似于邮政编码)，这些基序决定它们在细胞内的末端位置。细胞表膜蛋白是在内质网中合成的。其中许多蛋白质携带一组信号，如果它们不能形成正确折叠的蛋白质，就会导致它们被保留在细胞内(主要是内质网)。一些膜蛋白还携带另一组信号，帮助它们离开内质网并传递到细胞表面。人们认为，当蛋白质正确折叠时，它有能力隐藏导致它停留在内质网内的信号，并暴露有助于其运输到细胞内正确位置的信号。提出了一种一般的策略，在内质网中合成的蛋白质通过从内质网中萌发并与高尔基体融合的运输小泡(称为CoP II小泡)运输到高尔基体。如果这些囊泡中携带的蛋白质被错误折叠，并且具有未被覆盖的内质网保留信号/基序，则它们被放置在不同的运输囊泡(COPI囊泡)中并返回到内质网。如果蛋白质要到达细胞表面，它们必须隐藏它们的保留基序。保留基序的掩蔽是通过膜蛋白的正确折叠和与额外蛋白的相互作用来实现的。我们之前已经证明，内质网保留基序可以被一种名为14-3-3的蛋白质掩盖。到目前为止，一组十几种蛋白质被证明需要1433才能使它们走出内质网。这个项目将研究14-3-3帮助蛋白质到达其在细胞表面的正确位置的机制。许多问题仍有待回答，包括14-3-3是仅在内质网结合并掩蔽膜蛋白上的ER保留信号，还是它与膜蛋白一起一直移动到细胞表面。如果14-3-3真的一直飞到地表，它对那里有影响吗？或者，14-3-3的作用是使细胞表面膜蛋白更容易或更快地到达目的地？其他蛋白质是否也帮助膜蛋白进入细胞表面？由于14-3-3似乎有助于膜蛋白到达细胞表面，如果我们能够确定帮助14-3-3与这些蛋白相互作用的条件，我们就可以增加细胞表面的蛋白质数量。同样，我们也可以防止相互作用，减少到达表面的膜蛋白数量。由于膜蛋白对细胞的功能至关重要，我们可以通过一种机制来控制它们在细胞表面的数量，这将为改变健康和疾病中的细胞功能提供一种手段。

Cells respond to and communicate with their surroundings through the proteins (receptors, ion channels and transporters) they present on their surface. Receptors are embedded in the cell membrane and allow the cell to recognise and respond to specific signals which result in a cellular response. Ion channels and transporters also play a role in controlling the response of cells to stimuli by controlling the flow of ions into and out of the cell. The number and location of these membrane proteins on the cell surface can greatly affect the response of cells to external signals while mis-location of these critical cell surface proteins can give rise to conditions such as diabetes and cystic fibrosis. This project will investigate one mechanism by which cells control the number of membrane proteins on the cell surface. Cells have a number of ways of monitoring and controlling the number and type of membrane proteins on their surface. Proteins carry signal motifs (similar to postal codes) which determine their end location within the cell. Cell surface membrane proteins are synthesised in the endoplasmic reticulum (ER). Many of these proteins carry a set of signals which results in them being held within the cell (primarily the ER) if they fail to form correctly folded proteins. Some membrane proteins also carry another set of signals which aid their exit from the ER and delivery to the cell surface. It is thought that when a protein is folded correctly it has the ability to hide signals that result in it staying within the ER and expose signals that will aid its transport to the correct location within the cell. A general strategy is proposed, proteins synthesised in the ER are transported to the Golgi apparatus by transport vesicles (called COP II vesicles) which bud off from the ER and fuse with the Golgi apparatus. If the proteins carried in these vesicles are incorrectly folded and have an uncovered ER retention signal / motif they are then placed in different transport vesicles (COPI vesicles) and returned to the ER. If the proteins are destined to reach the cell surface they must conceal their retention motifs. Masking of retention motifs occurs through correct folding of the membrane proteins and interaction with additional proteins. We have previously demonstrated that ER retention motifs can be masked by a protein called 14-3-3. To date a group of more than a dozen proteins have been shown to require 14 3 3 to allow them move out of the ER. This project will examine the mechanism by which 14-3-3 helps proteins reach their correct location on the cell surface. Many questions remain to be answered, including does 14-3-3 bind to and mask the ER retention signals on membrane proteins only at the ER or does it travel with the membrane protein all the way to the cell surface. If 14-3-3 does travel all the way to the surface does it have an effect there? Or is the role of 14-3-3 to make it easier or faster for cell surface membrane proteins to reach thier destinations? Do other proteins also help membrane proteins make their way to the cell surface? As 14-3-3 appears to help membrane proteins get to the cell surface, if we can determine conditions which help 14-3-3 interact with these proteins we can increase the number of proteins on the cell surface. Similarly we could also prevent the interaction and reduce the number of membrane proteins reaching the surface. As membrane proteins are critical to the functioning of cells a mechanism by which we can control their number on the cell surface would provide a means to alter cell function in health and disease.