Cell polarity and phosphoinositide kinases in Wnt signalling

Wnt 信号传导中的细胞极性和磷酸肌醇激酶

• 批准号: BB/J015075/1
• 负责人: Jeremy Green
• 金额: $ 44.09万
• 依托单位: King's College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FJ015075%2F1
• 关键词: Cell; polarity; phosphoinositide; kinases; Wnt

A large proportion of colon cancers and a significant fraction of other cancers in humans are due to problems with a chemical signal known as the "wnt" protein. Wnt is sent by certain cells in the body to other cells normally to promote healthy growth and renewal. When these signals become excessive, cancer is the result. The Wnt signal, as well as being important in cancer, is also very important in normal tissues during embryo development. In the right places and times, Wnt triggers the formation of different kinds of tissues and their growth so that the body forms correctly. If we can understand how Wnt signalling works, we may improve cancer therapies and harness normal embryonic processes to heal and repair tissues affected by injury, disease and ageing. One complication of Wnt signalling is that Wnt can trigger three different kinds of response and it is not understood what determines which of these responses a cell will give. We recently found an enzyme, known as PI4KIIb, which, when depleted from frog embryos, causes the resulting tadpoles to have shorter, wider bodies and smaller heads. (We use frog embryos because they are available in large numbers and they and their cells are large and so easy to manipulate and observe.) The shorter, wider bodies we see are suggestive of effects on so-called non-canonical Wnt signalling (of which there are two types). The smaller heads are suggestive of effects of increased canonical Wnt-beta-catenin signalling. It is possible that PI4KIIb affects both, but a very similar enzyme, PI4KIIa, affects only canonical signalling, and depleting the latter from embryos increases canonical Wnt signalling and head size. Our first objective is to test directly whether the effects we see on the tadpole appearance really are due to the effects on the different Wnt pathways, or whether something else is going on. We will measure how much of each response occurs in tadpoles that have increased or decreased amounts of PI4KIIa and b. In a previous project, we found that the different Wnt pathway branches are regulated by different versions of another enzyme known as PAR-1. We have now found that PAR-1 acts on PI4KIIb, adding a phosphate group to a specific site on the latter. Moreover, we find that this site in PI4KIIb is needed for it to move to the cell's surface membrane. Others have shown that membrane association is critical for activity of PI4KIIb. We will test whether PAR-1a affects PI4KIIa in the same way as it does PI4KIIb by similarly mutating the target site and seeing whether the mutated protein is prevented from going to the cell membrane. We shall also test whether depleting different versions of PAR-1 affects the localisation of PI4KIIa and -b in embryos, with the idea that the different PAR-1 versions act on different PI4KII versions.Having established PI4KII pathway activities and localisation determinants, we shall see whether these two key properties of PI4KIIb are related: does PI4KII localisation affect its activity in Wnt signalling? We will do the same sets of pathway assays on the PI4KII versions and mutants and see whether the activity and localisation are correlated. Since PAR-1 is always found on the basal side of epithelial (layered) tissues, we will test the idea that the PAR-1/PI4KII system governs which cell responses occur when Wnt is presented to different sides of such tissues. This would be a major insight into the significance of the different pathways and their regulation by PAR-1 and show that in cancer therapy or regenerative medical approaches, it matters not just whether but also where in relation to the cells' polarity agents that mimic or inhibit Wnts are applied.

大部分的结肠癌和其他癌症都是由一种叫做“wnt”蛋白的化学信号引起的。Wnt通常由体内的某些细胞发送到其他细胞，以促进健康生长和更新。当这些信号变得过度时，癌症就是结果。Wnt信号在癌症中很重要，在胚胎发育过程中在正常组织中也很重要。在正确的地方和时间，Wnt触发不同类型组织的形成及其生长，以便身体正确形成。如果我们能够理解Wnt信号传导是如何工作的，我们可能会改善癌症治疗，并利用正常的胚胎过程来愈合和修复受损伤，疾病和衰老影响的组织。Wnt信号传导的一个复杂之处在于，Wnt可以触发三种不同类型的反应，并且还不清楚是什么决定了细胞将给出哪种反应。我们最近发现了一种被称为PI 4KIIb的酶，当这种酶从青蛙胚胎中耗尽时，会导致产生的蝌蚪具有更短，更宽的身体和更小的头部。(We使用青蛙胚胎，因为它们数量很多，它们和它们的细胞都很大，很容易操作和观察。我们看到的更短，更宽的身体暗示了对所谓的非经典Wnt信号传导的影响（其中有两种类型）。较小的头部提示增加的典型Wnt-β-连环蛋白信号传导的作用。PI 4KIIb可能影响两者，但一种非常相似的酶PI 4KIIa只影响典型信号传导，从胚胎中耗尽后者会增加典型Wnt信号传导和头部大小。我们的第一个目标是直接测试我们所看到的对蝌蚪外观的影响是否真的是由于对不同Wnt通路的影响，或者是否有其他原因。我们将测量在增加或减少PI 4 KIIa和B量的蝌蚪中发生的每种反应的程度。在之前的一个项目中，我们发现不同的Wnt通路分支受到另一种称为PAR-1的酶的不同版本的调节。我们现在已经发现PAR-1作用于PI 4KIIb，在后者的特定位点上添加磷酸基团。此外，我们发现PI 4KIIb中的这个位点是它移动到细胞表面膜所必需的。其他人已经表明，膜结合对于PI 4 KIIb的活性至关重要。我们将测试PAR-1a是否以与PI 4KIIb相同的方式影响PI 4KIIa，方法是类似地突变靶位点，并观察突变的蛋白质是否被阻止进入细胞膜。我们还将测试是否耗尽不同版本的PAR-1影响本地化的PI 4 KIIa和-B在胚胎中，与想法，不同的PAR-1版本的作用于不同的PI 4 KII版本。已建立PI 4 KII通路活动和本地化的决定因素，我们将看到这两个关键属性的PI 4 KII B是否相关：PI 4 KII本地化影响其在Wnt信号传导的活动？我们将在PI 4KII版本和突变体上进行相同的通路测定，看看活性和定位是否相关。由于PAR-1总是在上皮（分层）组织的基底侧发现，我们将测试PAR-1/PI 4KII系统控制当Wnt被呈递到这些组织的不同侧时发生哪些细胞反应的想法。这将是对不同途径的重要性及其通过PAR-1的调节的重要见解，并表明在癌症治疗或再生医学方法中，重要的不仅仅是是否而且与模拟或抑制Wnt的细胞极性试剂有关。

项目成果

PI4K2ß constrains non-canonical Wnt-PCP signalling and is localised by PAR-1 (MARK2/3) phosphorylation

PI4K2� 限制非典型 Wnt-PCP 信号传导并通过 PAR-1 (MARK2/3) 磷酸化进行定位

• DOI: 10.1101/660431
• 发表时间: 2019
• 作者: Tsang S