Activation of chimeric and full-length growth hormone receptors by growth hormone receptor monoclonal antibodies - A specific conformational change may be required for full-length receptor signaling
Activation of chimeric and full-length growth hormone receptors by growth hormone receptor monoclonal antibodies - A specific conformational change may be required for full-length receptor signaling
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DOI：
10.1074/jbc.273.9.5307
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发表时间：
1998-02-27
影响因子：
4.8
通讯作者：
Waters, MJ
中科院分区：
文献类型：
--
作者：
Rowlinson, SW;Behncken, SN;Waters, MJ
生长激素受体（GHR）的信号转导是通过生长激素（GH）诱导的两个GHR的二聚化形成三聚体来实现的，人们认为仅二聚化就足以信号转导。由于针对GHR细胞外结构域的单克隆抗体（mab）可诱导转染含有GHR细胞外结构域、纤维连接蛋白和小鼠粒细胞集束刺激因子受体的嵌合受体的FDC-P1细胞增殖，因此我们筛选了14种针对稳定表达全长人、兔或大鼠GHR的特征FDC-P1和BaF-B03细胞系的GHR单克隆抗体。或嵌合人GHR/粒细胞集落刺激因子受体，以及c-fos启动子的转激活和STAT激活。与嵌合受体，八个单克隆抗体能够诱导增殖，虽然没有抑制激素结合和激动剂活性之间的相关性。相比之下，没有单克隆抗体能够与全长GHR FDC-P1细胞系作为激动剂，尽管有9种单克隆抗体与GH竞争结合，在BaF-B03细胞系中观察到两种单克隆抗体（263和1C9）的微弱增殖反应，并且抗小鼠F(ab)(2)的添加导致hGHR BaF-B03细胞系的信号传导增加，达到mAb 263最高GH的28 +/- 4%。这些数据可能表明单克隆抗体正确二聚化全长GI-IR的能力相当严格。然而，mAb 263刺激突变体hGHR在结构域2的F‘-G’环中改变的能力几乎被消除，同时该mAb对受体的亲和力增加。由于F‘-G’环在GH结合时经历构象变化，并且是充分增殖信号传导所必需的，我们提出除了促进受体二聚化外，mAb 263可能诱导类似GH的受体构象的特异性变化，这是生物反应所必需的。
Signal transduction by the growth hormone receptor (GHR) occurs through growth hormone (GH)-induced dimerization of two GHRs to form a trimeric complex, It is thought that dimerization alone is sufficient for signaling, since monoclonal antibodies (mAbs) against the extracellular domain of the GHR elicit proliferation of FDC-P1 cells transfected with a chimeric receptor comprising the extracellular domain of the GHR and the fibronectin and cytoplasmic domains of the murine granulocyte colony-stimulating factor receptor, We have screened 14 GHR mAbs for proliferative activity against characterized FDC-P1 and BaF-B03 cell lines stably expressing the full-length human, rabbit, or rat GHR, or the chimeric human GHR/granulocyte colony-stimulating factor receptor, and for transactivation of the c-fos promoter and STAT activation. With the chimeric receptor, eight mAbs were able to elicit proliferation, although there was no correlation between inhibition of hormone binding and agonist activity. In contrast, no mAbs were able to act as agonists with the full-length GHR FDC-P1 cell lines, although nine competed with GH for binding, A weak proliferative response was observed in the BaF-B03 cell lines with two of the mAbs (263 and 1C9), and the addition of anti-mouse F(ab)(2) resulted in increased signaling in the hGHR BaF-B03 cell line to a plateau of 28 +/- 4% of the GH maximum for mAb 263. These data could indicate considerable stringency in the ability of mAbs to correctly dimerize the full-length GI-IR. However, the ability of mAb 263 to stimulate a mutant hGHR altered in the F'-G' loop of domain 2 was nearly abolished, concurrent with an increased affinity of this mAb for the receptor. Since the F'-G' loop undergoes a conformational change on GH binding and is necessary for full proliferative signaling, we propose that in addition to promoting receptor dimerization, mAb 263 may induce specific changes in receptor conformation similar to GH, which are required for the biological response.