Corticotropin-releasing hormone receptor (CRHR)1 and CRHR2 are both trafficking and signaling receptors for urocortin.
Corticotropin-releasing hormone receptor (CRHR)1 and CRHR2 are both trafficking and signaling receptors for urocortin.
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DOI：
10.1210/me.2005-0503
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发表时间：
2007-03
影响因子：
--
通讯作者：
H. Tu;A. Kastin;W. Pan
中科院分区：
文献类型：
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作者：
H. Tu;A. Kastin;W. Pan
尿皮质素是一种有效的饱腹感肽，其转运发生在小鼠的血脑屏障处。构成血脑屏障的脑微血管内皮细胞对尿皮质素的内吞作用是这种运输的限速步骤，但所涉及的细胞机制尚未完全阐明。本研究中分离的脑微血管中存在 CRH 受体 R1 和 R2，表明这两种亚型可能介导尿皮质素转运。通过人胚胎肾 293 细胞中的过表达研究测试了这两种受体在尿皮质素内吞作用和信号转导中的作用。两种受体均导致放射性标记的尿皮质素的结合和内吞作用显着增加。 CRHR1 介导的尿皮质素内吞作用被 astressin（两种 CRHR 的拮抗剂）阻断，而 CRHR2 介导的尿皮质素内吞作用也被 antisauvagine 30（选择性 CRHR2β 拮抗剂）阻断。氯丙嗪、菲律宾平和制霉菌素对尿皮质素内吞作用没有影响，表明网格蛋白或小凹膜微区没有显着参与。 CRHR1 和 CRHR2 都能介导配体诱导的 cAMP 产生增加，表明过度表达的受体具有生物活性。然而，毛喉素或二丁酰-cAMP 升高细胞内 cAMP 并没有显示出对尿皮质素的结合和内吞作用的急性调节。尽管在过表达 CRHR1 或 CRHR2 的细胞中内吞的尿皮质素在细胞内大量降解，但完整的尿皮质素可以在 1 小时的研究间隔期间被胞吐。我们得出的结论是，CRHR1 和 CRHR2 在这种有效肽的非网格蛋白、非小窝介导的内吞作用和细胞内信号转导中发挥促进作用。
Transport of urocortin, a potent satiety peptide, occurs at the blood-brain barrier of the mouse. Endocytosis of urocortin by the cerebral microvessel endothelial cells composing the blood-brain barrier is a rate-limiting step of this transport, but the cellular mechanisms involved have not been fully elucidated. The presence of both CRH receptors R1 and R2 in isolated cerebral microvessels shown in this study suggested that both subtypes might mediate urocortin transport. The roles of these two receptors in the endocytosis and signal transduction of urocortin were tested by overexpression studies in human embryonic kidney 293 cells. Both receptors led to a significant increase of binding and endocytosis of radiolabeled urocortin. CRHR1-mediated urocortin endocytosis was blocked by astressin (antagonist for both CRHRs), whereas CRHR2-mediated urocortin endocytosis was also blocked by antisauvagine 30 (selective CRHR2beta antagonist). Chlorpromazine, filipin, and nystatin had no effect on urocortin endocytosis, indicating the lack of significant involvement of clathrin or caveolae membrane microdomains. Both CRHR1 and CRHR2 were able to mediate the ligand-induced increase of cAMP production, suggesting that the overexpressed receptors were biologically active. Elevation of intracellular cAMP by forskolin or dibutyryl-cAMP, however, did not show acute modulation of the binding and endocytosis of urocortin. Despite the substantial intracellular degradation of endocytosed urocortin in cells overexpressing either CRHR1 or CRHR2, intact urocortin could be exocytosed during the 1-h study interval. We conclude that both CRHR1 and CRHR2 play a facilitatory role in the non-clathrin-, non-caveolae-mediated endocytosis and intracellular signal transduction of this potent peptide.