Volpicelli Laura A, Lah James J, Fang Guofu, Goldenring James R, Levey Allan I
Center for Neurodegenerative Disease and Department of Neurology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
J Neurosci. 2002 Nov 15;22(22):9776-84. doi: 10.1523/JNEUROSCI.22-22-09776.2002.
Agonist-induced internalization followed by subsequent return to the cell surface regulates G-protein-coupled receptor (GPCR) activity. Because the cellular responsiveness to ligand depends on the balance between receptor degradation and recycling, it is crucial to identify the molecules involved in GPCR recovery to the cell surface. In this study, we identify mechanisms involved in the recycling of the M4 subtype of muscarinic acetylcholine receptor. M4 is highly expressed in the CNS, plays a role in locomotor activity, and is a novel therapeutic target for neurologic and psychiatric disorders. Previous studies show that, after cholinergic stimulation, M4 internalizes from the cell surface to endosomes in cell culture and the rat brain. Here, we show that, after activation, M4 traffics to transferrin receptor- and Rab11a-positive perinuclear endosomes. Expression of the constitutively GDP-bound, inactive mutant Rab11aS25N inhibits M4 trafficking to recycling endosomes. Expression of the C-terminal tail of myosin Vb, a Rab11a effector, enhances M4 accumulation in perinuclear endosomes. Both Rab11aS25N and the myosin Vb tail impair M4 recycling. The results demonstrate that GPCR recycling is mediated through a discrete pathway using both Rab11a and myosin Vb.
激动剂诱导的内化作用随后返回细胞表面可调节G蛋白偶联受体（GPCR）的活性。由于细胞对配体的反应性取决于受体降解和再循环之间的平衡，因此识别参与GPCR恢复到细胞表面的分子至关重要。在本研究中，我们确定了毒蕈碱型乙酰胆碱受体M4亚型再循环所涉及的机制。M4在中枢神经系统中高度表达，在运动活动中起作用，并且是神经和精神疾病的新型治疗靶点。先前的研究表明，在胆碱能刺激后，M4在细胞培养物和大鼠脑中从细胞表面内化到内体中。在这里，我们表明，激活后，M4转运至转铁蛋白受体和Rab11a阳性的核周内体。组成型GDP结合的无活性突变体Rab11aS25N的表达抑制M4向内体再循环的转运。Rab11a效应器肌球蛋白Vb的C末端尾巴的表达增强了M4在核周内体中的积累。Rab11aS25N和肌球蛋白Vb尾巴均损害M4的再循环。结果表明，GPCR再循环是通过使用Rab11a和肌球蛋白Vb的离散途径介导的。
