T-cell interactions with second generation glass-supported lipid bilayers

在关于静息T细胞表面的分子组织和动力学的剩余问题的核心的关键蛋白质之一是Src型酪氨酸激酶，Lck，其通过磷酸化T细胞受体（TCR）来启动T细胞中的信号传导。Lck具有相当大的内在兴趣，首先是因为其活性需要受到限制以防止不受控制的T细胞活化，其次是因为它是一种不寻常的表面组分，因为它通过两种类型的脂质型锚定物即肉豆蔻酰基和棕榈酰基与膜缔合，并且尚不清楚这如何或是否影响该激酶的分布和/或功能。在TCR的情况下，已经提出受体连接诱导受体周围脂质环境的变化，这促进下游信号传导分子（包括激酶）的进入。感兴趣的第三点是，与其他Src激酶相比，Lck与辅助受体CD 4相关联，CD 4是一种整合的膜蛋白。这是可能的，脂质微环境和动态的CD 4结合和未结合LCK diffusion different.We建议表征的组织和相互作用的动态的脂质锚定蛋白在T细胞表面，即豆蔻酰化和棕榈酰化LCK，无论是在不同的脂质，并在比较其他组件的T细胞的信号机制，包括CD 4，使用STED（-FCS）显微镜。我们希望这些新的实验能够以前所未有的细节突出T细胞表面关键信号蛋白的组织，并为理解受体触发建立一个关键框架。

One of the key proteins at the heart of the remaining questions on the molecular organization and dynamics of the resting T-cell surface is the Src-type tyrosine kinase, Lck, which initiates signalling in T-cells by phosphorylating the T-cell receptor (TCR). Lck is of considerable intrinsic interest firstly because its activity needs to be constrained in order to prevent uncontrolled T-cell activation, and secondly because it is an unusual surface component insofar as it associates with the membrane via two types of lipid-type anchors i.e. myristoyl and palmitoyl groups, and it is unclear how or whether this affects the distribution and/or function of this kinase. In the case of the TCR, it has been suggested that receptor ligation induces changes in the lipid environment around the receptor, which facilitates the access of downstream signalling molecules, including kinases. A third point of interest is that, in contrast to other Src kinases, Lck associates with the co-receptor CD4, which is an integral membrane protein. It is possible that the lipid micro-environment and dynamics of CD4-bound and -unbound Lck diffusion differ.We propose to characterize the organization and interaction dynamics of a lipid-anchored protein at the T-cell surface, i.e. myristoylated and palmitoylated Lck, both in relation to different lipids, and in comparison to other components of the signalling machinery of the T-cell including CD4, using STED(-FCS) microscopy. We expect these novel experiments to highlight, in thus far unprecedented detail, the organization of key signalling proteins at the T-cell surface, and to create a critical framework for understanding receptor triggering.