Agonist-Activated Bombyx Corazonin Receptor Is Internalized via an Arrestin-Dependent and Clathrin-Independent Pathway.
Agonist-Activated Bombyx Corazonin Receptor Is Internalized via an Arrestin-Dependent and Clathrin-Independent Pathway.
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DOI：
10.1021/acs.biochem.6b00250
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发表时间：
2016-07
期刊：
影响因子：
2.9
通讯作者：
Jingwen Yang;Zhangfei Shen;Xue一hua Jiang;Huipeng Yang;Haishan Huang;Lili Jin;Yajie Chen;Liangen Shi;N. Zhou
中科院分区：
文献类型：
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作者：
Jingwen Yang;Zhangfei Shen;Xue一hua Jiang;Huipeng Yang;Haishan Huang;Lili Jin;Yajie Chen;Liangen Shi;N. Zhou
激动剂诱导的内化在G蛋白偶联受体信号传导的程度和持续时间的严格调节中起关键作用。以前，我们已经表明，BmCrzR的corazonin受体（BmCrzR）激活Gαq和Gα s依赖的信号级联。然而，参与调节BmCrzR的内化和脱敏的分子机制仍有待阐明。这里，用于表达在C-末端融合有增强型绿色荧光蛋白（EGFP）的BmCrzR的载体用于进一步表征BmCrzR内化。我们发现在HEK-293和BmN细胞中异源表达的BmCrzR响应于激动剂激发，以浓度和时间依赖性方式通过β-抑制蛋白（Kurtz）依赖性和网格蛋白非依赖性途径从质膜迅速内化到细胞质中。虽然大多数内化的受体通过早期内体再循环到细胞表面，但一些其他受体被转运到溶酶体进行降解。使用RNA干扰的分析显示，GRK 2和GRK 5基本上参与BmCrzR磷酸化和内化的调节。进一步的研究表明，所鉴定的Ser/Thr残基簇（（411）TSS（413））负责GRK介导的磷酸化和内化。这是第一次详细调查的内在化和贩运的corazonin受体。
Agonist-induced internalization plays a key role in the tight regulation of the extent and duration of G protein-coupled receptor signaling. Previously, we have shown that the Bombyx corazonin receptor (BmCrzR) activates both Gαq- and Gαs-dependent signaling cascades. However, the molecular mechanisms involved in the regulation of the internalization and desensitization of BmCrzR remain to be elucidated. Here, vectors for expressing BmCrzR fused with enhanced green fluorescent protein (EGFP) at the C-terminal end were used to further characterize BmCrzR internalization. We found that the BmCrzR heterologously expressed in HEK-293 and BmN cells was rapidly internalized from the plasma membrane into the cytoplasm in a concentration- and time-dependent manner via a β-arrestin (Kurtz)-dependent and clathrin-independent pathway in response to agonist challenge. While most of the internalized receptors were recycled to the cell surface via early endosomes, some others were transported to lysosomes for degradation. Assays using RNA interference revealed that both GRK2 and GRK5 were essentially involved in the regulation of BmCrzR phosphorylation and internalization. Further investigations indicated that the identified cluster of Ser/Thr residues ((411)TSS(413)) was responsible for GRK-mediated phosphorylation and internalization. This is the first detailed investigation of the internalization and trafficking of Bombyx corazonin receptors.