Molecular Recognition Studies of the Interleukin-7 Pathway

描述（由申请人提供）：白细胞介素-7（IL-7）信号通路在细胞外基质建模以及B和T细胞的发育和稳态中起重要作用。异常的IL-7信号转导与严重的联合免疫缺陷、自身免疫性疾病和癌症有关。当可溶性IL-7与两种细胞表面细胞因子受体（白细胞介素-7受体（IL-7 R）和共同γ链）相互作用时，触发IL-7信号。在上一个资助期，我们确定了结构，能量和细胞生物学 IL-7与IL-7 R相互作用的特征。在本基金资助期内，我们将深入研究IL-7与糖胺聚糖（如肝素/硫酸肝素）以及IL-7和IL-7 R共同的γ链受体相互作用的分子机制。对这些相互作用的初步研究揭示了意外的非活化复合物的形成，所述非活化复合物是IL-7信号传导所特有的或对于其他常见的γ链家族成员未鉴定的。具体地，糖胺聚糖结合诱导IL-7的二聚体形成，并且共同的γ-链受体可以不依赖于IL-7 R而与IL-7结合，并且不依赖于IL-7而与IL-7 R结合.进一步的研究将包括非活化IL-7/GAG、IL-7/共同γ链、IL-7 R/共同γ链复合物以及活化IL-7/IL-7 R/共同γ链三元复合物的三维X射线晶体学结构。这些表位的结合决定簇也将使用定点扫描诱变和各种生物物理方法（包括等温滴定量热法和表面等离子体共振）来确定。非活化和活化复合物的结构和生物物理研究最终将用于开发解释正常和异常IL-7信号传导机制的模型，并产生治疗与IL-7信号传导受损相关的多种疾病的新型蛋白质试剂和治疗剂。

DESCRIPTION (provided by applicant): The interleukin-7 (IL-7) signaling pathway plays essential roles in modeling the extracellular matrix and the development and homeostasis of B and T cells. Aberrant IL-7 signaling has been implicated in severe combined immunodeficiency, autoimmune conditions, and cancers. The IL-7 signal is triggered when soluble IL-7 interacts with two cell surface cytokine receptors, interleukin-7 receptor (IL-7R) and the common gamma- chain. In the previous funding period, we determined the structural, energetic, and cell biological features of the interaction of IL-7 with IL-7R. In this funding period, we will dissect the moleculr mechanisms of the interactions of IL-7 with glycosaminoglycans (e.g. heparin/heparin sulfate) and the common gamma-chain receptor with IL-7 and IL-7R. Preliminary studies of these interactions reveal the formation of unexpected non- activating complexes, either unique to IL-7 signaling or unidentified for the other common gamma-chain family members. Specifically, glycosaminoglycan binding induces dimer formation of IL-7, and common gamma- chain receptor can bind to IL-7 independent of IL-7R and to IL-7R independent of IL-7. Further studies will include three-dimensional x-ray crystallographic structures of the non-activating IL-7/GAG, IL-7/common gamma-chain, IL-7R/common gamma-chain complexes, as well as the activating IL-7/IL-7R/common gamma- chain ternary complex. The binding determinants of these epitopes also will be determined using site-directed scanning mutagenesis and various biophysical methods, including isothermal titration calorimetry and surface plasmon resonance. The structural and biophysical studies of non-activating and activating complexes ultimately will be used to develop models that explain normal and aberrant IL-7 signaling mechanisms and generate novel protein reagents and therapeutics that treat the numerous diseases associated with impaired IL-7 signaling.