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>The adaptor protein EBP50 is important for localization of the protein kinase A–Ezrin complex in T-cells and the immunomodulating effect of cAMP
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The adaptor protein EBP50 is important for localization of the protein kinase A–Ezrin complex in T-cells and the immunomodulating effect of cAMP
pWe recently reported that the dual-specificity AKAP (A-kinaseanchoring protein) Ezrin targets type I PKA (protein kinase A) to the vicinity of the TCR (T-cell receptor) in T-cells and, together with PAG (phosphoprotein associated with glycosphingolipid-enriched membrane microdomains) and EBP50 [ERM (Ezrin/Radixin/Moesin)-binding phosphoprotein 50], forms a scaffold that positions PKA close to its substrate, Csk (C-terminal Src kinase). This complex is important for controlling the activation state of T-cells. Ezrin binds the adaptor protein EBP50, which again contacts PAG. In the present study, we show that Ezrin and EBP50 interact with high affinity (iK/isubD/sub=58±7 nM). A peptide corresponding to the EB (Ezrin-binding) region in EBP50 (EBP50subpep/sub) was used to further characterize the binding kinetics and compete the Ezrin–EBP50 interaction by various methods iin vitro/i. Importantly, loading T-cells with EBP50subpep/sub delocalized Ezrin, but not EBP50. Furthermore, disruption of this complex interfered with cAMP modulation of T-cell activation, which is seen as a reversal of cAMP-mediated inhibition of IL-2 (interleukin 2) production, demonstrating an important role of EBP50 in this complex. In summary, both the biochemical and functional data indicate that targeting the Ezrin–EBP interaction could be a novel and potent strategy for immunomodulation./p
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机译：>我们最近报道说，双重特异性AKAP（A激酶锚定蛋白）Ezrin将I型PKA（蛋白激酶A）靶向到T细胞中TCR（T细胞受体）附近，并与PAG一起使用（与糖鞘脂富集的膜微结构域相关的磷蛋白）和EBP50 [ERM（Ezrin / Radixin / Moesin）结合磷蛋白50]形成一个支架，将PKA置于其底物Csk（C末端Src激酶）附近。这种复合物对于控制T细胞的激活状态很重要。 Ezrin与衔接子蛋白EBP50结合，后者再次与PAG接触。在本研究中，我们显示Ezrin和EBP50以高亲和力相互作用（ K D = 58±7nM）。通过多种方法体外，使用对应于EBP50中EB（Ezrin结合）区域的肽（EBP50 pep ）进一步表征结合动力学并竞争Ezrin-EBP50相互作用。我>。重要的是，用EBP50 pep 离域的Ezrin（而不是EBP50）加载T细胞。此外，这种复合物的破坏会干扰cAMP对T细胞活化的调节，这被看作是cAMP介导的IL-2（白介素2）产生的抑制作用的逆转，证明了EBP50在该复合物中的重要作用。总之，生化和功能数据均表明，靶向Ezrin-EBP相互作用可能是一种新颖而有效的免疫调节策略。
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