Molecular Recognition Studies of the Interleukin-7 Pathway

描述(申请人提供)：白介素7(IL-7)信号通路在模拟细胞外基质以及B和T细胞的发育和动态平衡方面起着至关重要的作用。IL-7信号的异常与严重的联合免疫缺陷、自身免疫状况和癌症有关。当可溶性IL-7与两种细胞表面细胞因子受体--白介素7受体(IL-7R)和共同的伽马链相互作用时，IL-7信号被触发。在之前的资助期间，我们确定了结构、能量和细胞生物学 IL-7与IL-7R相互作用的特点在这一资助期间，我们将剖析IL-7与糖胺聚糖(例如肝素/硫酸肝素)相互作用的分子机制，以及常见的伽玛链受体与IL-7和IL-7R的相互作用。对这些相互作用的初步研究揭示了意想不到的非激活复合体的形成，这些复合体要么是IL-7信号所特有的，要么是其他常见的伽马链家族成员的未知复合体。具体来说，糖胺多糖结合诱导IL-7二聚体的形成，共同的伽马链受体可以独立于IL-7R与IL-7结合，也可独立于IL-7与IL-7R结合。进一步的研究将包括非激活的IL-7/GAG、IL-7/公共伽马链、IL-7R/公共伽马链复合体以及激活的IL-7/IL-7R/公共伽马链三元复合体的三维X射线晶体结构。这些表位的结合决定因素也将使用定点扫描突变和各种生物物理方法来确定，包括等温滴定量热法和表面等离子体共振。非激活和激活复合体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信号机制的模型，并产生新的蛋白质试剂和疗法，用于治疗与IL-7信号受损相关的许多疾病。

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.