Characterization of bombesin receptors in a rat pituitary cell line.
Characterization of bombesin receptors in a rat pituitary cell line.
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DOI：
10.1016/s0021-9258(18)32209-9
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发表时间：
1983-06
期刊：
影响因子：
--
通讯作者：
J. Westendorf;A. Schonbrunn
中科院分区：
文献类型：
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作者：
J. Westendorf;A. Schonbrunn
Bombesin 是一种十四肽，可刺激大鼠和人类以及 GH4C1 细胞培养物中催乳素的分泌，GH4C1 细胞是大鼠垂体肿瘤细胞的克隆株。我们利用[125I-Tyr4]铃蟾肽来鉴定和表征 GH4C1 细胞中的特异性高亲和力受体。对 4°C 平衡结合数据的 Scatchard 分析表明存在一类铃蟾肽非相互作用结合位点（RT = 3600 +/- 500 位点/细胞）。平衡解离常数值 (Kd = 1.2 +/- 0.4 nM) 与铃蟾肽刺激催乳素释放的 ED50 (0.5 nM) 非常一致。 37°C 稳定状态下的[125I-Tyr4]铃蟾肽结合可通过增加未标记铃蟾肽的浓度以剂量依赖性方式受到抑制，ID50 = 1.4 +/- 0.2 nM。然而，[125I-Tyr4]铃蟾肽的结合不受 100 nM 促甲状腺素释放激素、血管活性肠肽、表皮生长因子或生长抑素的抑制。因此，[125I-Tyr4]铃蟾肽结合的受体不同于调节 GH4C1 细胞激素分泌的其他肽的受体。高亲和力结合的类似物特异性表明铃蟾肽受体识别分子中的COOH末端八肽序列。在测试的五种垂体细胞株中，两种含有[125I-Tyr4]铃蟾肽（GH4C1和GH3）饱和结合位点的垂体细胞株先前已被证明对铃蟾肽有反应，激素分泌增加，而三种缺乏受体（GC、F4C1和AtT20/D16v）则无反应。因此，[125I-Tyr4]铃蟾肽结合位点似乎对于铃蟾肽的生物作用是必需的。对受体结合肽的加工和代谢的检查表明，在 4°C 时，[125I-Tyr4] 铃蟾肽与 GH4C1 细胞表面的受体结合。在 37 摄氏度时，受体结合肽迅速内化，随后在溶酶体中降解。总之，我们首次表征了特定的、高亲和力的垂体铃蟾肽受体，这些受体对于铃蟾肽的生物作用是必需的。
Bombesin is a tetradecapeptide which stimulates prolactin secretion in rats and man and in cultures of GH4C1 cells, a clonal strain of rat pituitary tumor cells. We have utilized [125I-Tyr4]bombesin to identify and characterize specific high affinity receptors in GH4C1 cells. Scatchard analysis of equilibrium binding data at 4 degrees C indicated the presence of a single class of non-interacting binding sites for bombesin (RT = 3600 +/- 500 sites/cell). The value for the equilibrium dissociation constant (Kd = 1.2 +/- 0.4 nM) agreed closely with the ED50 (0.5 nM) for bombesin stimulation of prolactin release. [125I-Tyr4]Bombesin binding at steady state at 37 degrees C was inhibited by increasing concentrations of unlabeled bombesin in a dose-dependent manner, with an ID50 = 1.4 +/- 0.2 nM. However, binding of [125I-Tyr4] bombesin was not inhibited by 100 nM thyrotropin-releasing hormone, vasoactive intestinal peptide, epidermal growth factor, or somatostatin. Therefore, [125I-Tyr4]bombesin binds to a receptor distinct from the receptors for other peptides which regulate hormone secretion by GH4C1 cells. The analog specificity for high affinity binding showed that the receptors for bombesin recognize the COOH-terminal octapeptide sequence in the molecule. Among five pituitary cell strains tested, two which contained saturable binding sites for [125I-Tyr4]bombesin (GH4C1 and GH3) had previously been shown to respond to bombesin with increased hormone secretion, whereas three which lacked receptors (GC, F4C1, and AtT20/D16v) were unresponsive. Therefore, the [125I-Tyr4]bombesin binding sites appear to be necessary for the biological actions of bombesin. Examination of the processing and metabolism of receptor-bound peptide demonstrated that at 4 degrees C [125I-Tyr4]bombesin binds to receptors on the surface of GH4C1 cells. At 37 degrees C, receptor-bound peptide is rapidly internalized and subsequently degraded in lysosomes. In summary, we have characterized for the first time specific, high affinity pituitary bombesin receptors which are necessary for the biological action of bombesin.