Gβγ subunit activation promotes dopamine efflux through the dopamine transporter.
Gβγ subunit activation promotes dopamine efflux through the dopamine transporter.
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DOI：
10.1038/mp.2017.176
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发表时间：
2017-12
影响因子：
11
通讯作者：
Torres GE
中科院分区：
文献类型：
--
作者：
Garcia-Olivares J;Baust T;Harris S;Hamilton P;Galli A;Amara SG;Torres GE
多巴胺转运蛋白（DAT）是脑内多巴胺（DA）稳态的重要调节因子，控制DA信号的强度和持续时间。DAT是精神兴奋剂（如可卡因和安非他明）的作用靶点，在神经精神障碍（包括注意力缺陷多动障碍和药物成瘾）中起着重要作用。因此，彻底了解调节DAT功能的机制对于开发治疗DA相关脑疾病的临床干预措施是必要的。先前的研究已经揭示了大量的蛋白质-蛋白质相互作用影响DAT细胞定位和活性，这表明DA稳态的微调涉及多种机制。我们最近报道了G-蛋白β-γ（Gβγ）亚基直接与DAT结合并降低DA清除率。我们发现Gβγ通过DAT诱导DA的释放。特别是，在异源细胞和培养的原代多巴胺能神经元中，Gβγ结合/激活肽mSIRK通过DAT增加DA流出。添加Gβγ抑制剂gallein或DAT抑制剂可防止这种效应。DAT羧基端的582 - 596位残基被确定为Gβγ的主要结合位点。含有DAT的Gβγ相互作用结构域的达特肽阻断mSIRK诱导DA流出的能力，这与Gβγ与转运蛋白的直接相互作用一致。最后，G蛋白偶联受体毒蕈碱M5 R的激活通过Gβγ依赖性机制导致DAT介导的DA外排。总的来说，我们的数据表明，Gβγ与DAT相互作用，以促进DA流出。这一新的机制可能对脑DA稳态的调节具有重要意义。
The dopamine transporter (DAT) is an important regulator of brain dopamine (DA) homeostasis, controlling the intensity and duration of DA signaling. DAT is the target for psychostimulants—like cocaine and amphetamine—and plays an important role in neuropsychiatric disorders, including attention-deficit hyperactivity disorder and drug addiction. Thus, a thorough understanding of the mechanisms that regulate DAT function is necessary for the development of clinical interventions to treat DA-related brain disorders. Previous studies have revealed a plethora of protein–protein interactions influencing DAT cellular localization and activity, suggesting that the fine-tuning of DA homeostasis involves multiple mechanisms. We recently reported that G-protein beta-gamma (Gβγ) subunits bind directly to DAT and decrease DA clearance. Here we show that Gβγ induces the release of DA through DAT. Specifically, a Gβγ-binding/activating peptide, mSIRK, increases DA efflux through DAT in heterologous cells and primary dopaminergic neurons in culture. Addition of the Gβγ inhibitor gallein or DAT inhibitors prevents this effect. Residues 582 to 596 in the DAT carboxy terminus were identified as the primary binding site of Gβγ. A TAT peptide containing the Gβγ-interacting domain of DAT blocked the ability of mSIRK to induce DA efflux, consistent with a direct interaction of Gβγ with the transporter. Finally, activation of a G-protein-coupled receptor, the muscarinic M5R, results in DAT-mediated DA efflux through a Gβγ-dependent mechanism. Collectively, our data show that Gβγ interacts with DAT to promote DA efflux. This novel mechanism may have important implications in the regulation of brain DA homeostasis.
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