NEGATIVE SIGNALLING BY KILLER CELL INHIBITORY RECEPTORS

人类自然杀伤(NK)细胞表现出一种独特的激活机制，该机制主要受抑制性受体的调控，其中包括杀伤细胞抑制性受体(KIR)。KIR是主要的NK细胞受体，与经典的主要组织相容性复合体I类分子(MHC-I)结合，传递抑制NK细胞活化的负信号。KIR负信号的一个组成部分是蛋白酪氨酸磷酸酶SHP-1在受体细胞质区域内重新募集到磷酸化的酪氨酸残基。SHP-1如何发挥抑制NK细胞激活的功能，以及其他信号效应分子是否也在这一过程中发挥作用，仍有待确定。我们最近发现，蛋白酪氨酸激酶(PTK)，CSK，也可以与酪氨酸磷酸化的KIR结合。KIR对CSK的招募为我们未来的研究提供了一个令人兴奋的基础，因为这种PTK具有独特的能力，可以磷酸化Src家族受体近端PTK上的调节酪氨酸残基，并终止它们的活性。这种新的负信号机制尚未被确定为任何抑制性受体，我们假设CSK募集为KIR提供了在早期阶段抑制蛋白酪氨酸磷酸化介导的激活事件的能力。更多数据支持我们的假设，即KIR上两个单独的酪氨酸磷酸化位点的磷酸化状态直接调节效应器酶结合谱和功能结果。我们建议用三个特定的目标来验证我们的假设：(1)从生物化学的角度研究CSK在KIR中的募集及其对Src家族的PTK的影响；(2)确定CSK募集对KIR抑制NK细胞激活的功能贡献；(3)表征单个KIR酪氨酸在抑制性信号反应中的作用。这些研究将有助于我们理解KIR介导的负向信号的机制。更好地了解主要调节NK细胞激活的机制最终将有助于开发治疗方法，以便在癌症、病毒感染和预防组织移植排斥反应的治疗中有益地操纵它们的功能。

Human natural killer (NK) cells exhibit a unique activation mechanism that is dominantly regulated by inhibitory receptors, including Killer Cell Inhibitory Receptors (KIR). KIR are the major NK cell receptors that bind classical major histocompatibility complex class I molecules (MHC-I) and transduce negative signals that restrain NK cell activation. A component of KIR negative signalling is recruitment of the protein tyrosine phosphatase SHP-1 to phosphorylated tyrosine residues within the receptor cytoplasmic domain. How SHP-1 functions to inhibit NK cell activation and whether additional signalling effector molecules also play roles in this process remain to be established. We have recently discovered that the protein tyrosine kinase (PTK), Csk, can also bind to tyrosine phosphorylated KIR. Csk recruitment by KIR provides an exciting basis for our future studies, since this PTK has the unique capacity to phosphorylate the regulatory tyrosine residues on receptor proximal PTKs of the Src family and terminate their activity. Such a novel negative signalling mechanism has not yet been identified for any inhibitory receptor, and we hypothesize that Csk recruitment provides KIR with capacity to dampen protein tyrosine phosphorylation-mediated activation events at an early stage. Additional data supports our hypothesis that phosphorylation status of the two individual tyrosine phosphorylation sites on KIR directly regulates effector enzyme binding profiles and functional outcomes. We propose to test our hypotheses with three specific aims: (1) to biochemically characterize Csk recruitment to KIR and its impact on PTKs of the Src family, (2) to determine the functional contributions of Csk recruitment on KIR inhibition of NK cell activation, and (3) to characterize the roles of the individual KIR tyrosines on inhibitory signalling responses. These studies will contribute substantially to our understanding of the mechanisms of KIR-mediated negative signalling. A better understanding of the mechanisms that dominantly regulate NK cell activation should ultimately allow for the development of therapeutic methods to beneficially manipulate their functions in therapies for cancer, viral infection, and prevention of tissue transplant rejection.