Growth Hormone Receptor Dimerization & Disulfide Linkage

性状（申请人提供）：生长激素（GH）主要来源于脑垂体前叶，以内分泌方式调节脊椎动物的出生后生长和代谢。最近的研究还表明，自分泌源性GH和完整的GH轴在动物癌症的形成和行为中的作用。GH受体（GHR）是细胞表面糖蛋白细胞因子受体超家族成员，其在其胞外结构域（ECD）中结合GH并通过其胞内结构域（ICD）与JAK 2酪氨酸激酶的相互作用激活信号传导。我们最近对调节GHR可用性和活化的机制的研究揭示了令人兴奋的见解，包括：1）JAK 2缔合影响GHR表面呈递、稳定性和运输; 2）降低人T47 D乳腺癌细胞中的催乳素受体（PRLR）水平增加GHR丰度和GH敏感性; 3）新型的构象敏感性抗GHR ECD抗体可能是有用的GH拮抗剂。我们假设：1）JAK 2表达水平和PRLR表达水平强烈影响GH反应性; 2）GH诱导的GHR构象变化是GH信号传导的基础，是治疗干预的潜在靶点。我们的具体目标是：1.确定JAK 2和PRLR调节GHR加工、细胞表面稳定性和下调的机制。我们将研究GHR预二聚化如何影响JAK 2对GHR运输的调节以及GHR ICD酪氨酸残基对GH非依赖性和GH依赖性GHR运输的作用。将检查PRLR表达对乳腺癌细胞中GHR可用性、GHR-JAK 2关联和GH作用的影响。2.测定构象特异性抑制性抗GHR ECD单克隆抗体的体内效力。我们将在信号传导研究中表征两种抗GHR抗体和Fab片段的抑制特性，还评估PRLR表达的影响。我们将确定这些试剂拮抗GH信号传导和癌症外植体生长的体内功效。这些研究探讨了GH敏感性的重要决定因素。完成将揭示调节细胞表面GHR可用性和命运的机制，以及调节GH敏感性的治疗相关工具。

DESCRIPTION (provided by applicant): Growth hormone (GH), derived largely from the anterior pituitary, regulates postnatal growth and metabolism in vertebrates in an endocrine fashion. Recent studies also suggest roles for autocrine-derived GH and for an intact GH axis in formation and behavior of cancers in animals. GH receptor (GHR) is a cell surface glycoprotein cytokine receptor superfamily member that binds GH in its extracellular domain (ECD) and activates signaling via its intracellular domain's (ICD) interaction with the JAK2 tyrosine kinase. Our recent studies of mechanisms regulating GHR availability and activation reveal exciting insights, including: 1) JAK2 association influences GHR surface presentation, stability, and trafficking; 2) reducing prolactin receptor (PRLR) levels in human T47D breast cancer cells augments GHR abundance and GH sensitivity; 3) novel, conformationally-sensitive anti-GHR ECD antibodies may be useful GH antagonists. We hypothesize: 1) JAK2 expression levels and PRLR expression levels strongly influence GH responsiveness; 2) GH-induced GHR conformational changes that underlie GH signaling are potential targets for therapeutic intervention. Our specific aims are: 1. Determine mechanisms by which JAK2 and PRLR regulate GHR processing, cel surface stability, and downregulation. We will study how GHR predimerization impacts JAK2's modulation of GHR trafficking and the role of GHR ICD tyrosine residues on GH-independent and GH-dependent GHR trafficking. Effects of PRLR expression on GHR availability, GHR-JAK2 association, and GH actions in breast cancer cells will be examined. 2. Determine in vivo efficacy of conformation-specific inhibitory anti-GHR ECD monoclonal antibodies. We will characterize inhibitory properties of two anti-GHR antibodies and Fab fragments in signaling studies, also assessing impact of PRLR expression. We will determine in vivo efficacy of these reagents to antagonize GH signaling and cancer explant growth. These studies probe important determinants of GH sensitivity. Completion will reveal mechanisms regulating cell surface GHR availability and fate and therapeutically relevant tools to modulate GH sensitivity.