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ADP-ribosylation factor 6 regulates mu-opioid receptor trafficking and signaling via activation of phospholipase D2

机译:ADP-核糖基化因子6通过激活磷脂酶D2调节μ阿片受体的转运和信号传导

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Endocytosis of the mu-opioid receptor (MOPr) has been shown to play a protective role against the development of tolerance to opioid drugs by facilitating receptor reactivation and recycling. it has been further demonstrated, that the opioid-mediated and ADP-ribosylation factor (ARF)-dependent activation of phospholipase D2 (PLD2) is a prerequisite for MOPr endocytosis. In this study. we investigated which particular ARF protein is involved in opioid-mediated PLD2 activation and what are the mechanisms of ARF function in MOPr trafficking and signaling. By coexpressing the MOPr and dominant negative or constitutively active ARF mutants in human embryonic kidney (HEK) 293 cells and primary cultured cortical neurons as well as by using siRNA technology, we identified the ARF6 protein to be involved in the regulation of MOPr endocytosis. We also found that expression of an effector domain mutant of ARF6, which is incapable of activating PLD, blocked agonist-induced endocytosis suggesting that ARF6 function in MOPr trafficking is PLD2-mediated. Analogously, opioid-mediated activation of PLD2 is blocked in the presence of dominant negative ARF6 mutants. Finally, we also showed that ARF6 protein influences the recycling/reactivation of internalized MOPr and thus modulates agonist-induced MOPr desensitization. Together, these results provide evidence that ARF6 protein regulates MOPr trafficking and signaling via PLD2 activation and hence affects the development of opioid receptor desensitization and tolerance.
机译:业已证实,μ阿片受体(MOPr)的内吞作用通过促进受体的再活化和再循环,在抵抗阿片类药物耐受性的发展中起保护作用。进一步证明,阿片介导的和ADP-核糖基化因子(ARF)依赖性的磷脂酶D2(PLD2)活化是MOPr内吞的先决条件。在这个研究中。我们研究了阿片类药物介导的PLD2激活涉及哪种特定的ARF蛋白,以及MOPr转运和信号传导中ARF功能的机制是什么。通过在人类胚胎肾(HEK)293细胞和原代培养的皮层神经元中共表达MOPr和显性负性或组成性活性ARF突变体,以及使用siRNA技术,我们确定了ARF6蛋白参与MOPr内吞作用的调节。我们还发现,不能激活PLD的ARF6效应域突变体的表达阻断了激动剂诱导的内吞作用,提示ARF6在MOPr转运中的功能是PLD2介导的。类似地,在显性负性ARF6突变体的存在下,阿片样物质介导的PLD2的激活被阻断。最后,我们还表明,ARF6蛋白影响内在化MOPr的回收/再激活,从而调节激动剂诱导的MOPr脱敏。在一起,这些结果提供了证据,证明ARF6蛋白通过PLD2激活调节MOPr的运输和信号传导,因此影响阿片受体脱敏和耐受性的发展。

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