MDR1 function is sensitive to the phosphorylation state of myosin regulatory light chain
MDR1 function is sensitive to the phosphorylation state of myosin regulatory light chain
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DOI：
10.1016/j.bbrc.2010.05.084
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发表时间：
2010-07-16
影响因子：
3.1
通讯作者：
Ishmael, Jane E.
中科院分区：
文献类型：
--
作者：
Bajaj, Gaurav;Rodriguez-Proteau, Rosita;Ishmael, Jane E.
多药耐药蛋白1（MDR 1）由两个同源的半部分组成，由细胞内连接区隔开。已报道该接头可结合肌球蛋白调节轻链（RLC），但尚不清楚这如何在肌球蛋白II复合物中发生。我们表征了MDR 1-RLC相互作用，并确定结合通过RLC的氨基末端发生，RLC是一个通常结合肌球蛋白重链的结构域。MDR 1-RLC相互作用对轻链的磷酸化状态敏感，因为肌球蛋白轻链激酶（MLCK）的磷酸化导致体外结合丧失。我们使用ML-7，MLCK的特异性抑制剂，研究在完整细胞中破坏RLC磷酸化的功能后果。用ML-7预处理稳定表达MDR 1的极化Madin-Darby犬肾细胞，可显著增加[H-3]-地高辛（一种经典的MDR 1底物）的顶端至基底通透性，并相应降低[H-3]-地高辛的外排率（3倍; p < 0.01）。总之，这些数据表明，MDR 1介导的运输[H-3]-地高辛可以通过药理学操纵的肌球蛋白RLC调制，但直接MDR 1-RLC相互作用是非典型的，不能解释的肌球蛋白II全酶的结构。(C)2010年爱思唯尔公司All rights reserved.
Multiple drug resistance protein 1 (MDR1) is composed of two homologous halves separated by an intracellular linker region. The linker has been reported to bind myosin regulatory light chain (RLC), but it is not clear how this can occur in the context of a myosin II complex. We characterized MDR1-RLC interactions and determined that binding occurs via the amino terminal of the RLC, a domain that typically binds myosin heavy chain. MDR1-RLC interactions were sensitive to the phosphorylation state of the light chain in that phosphorylation by myosin light chain kinase (MLCK) resulted in a loss of binding in vitro. We used ML-7, a specific inhibitor of MLCK, to study the functional consequences of disrupting RLC phosphorylation in intact cells. Pretreatment of polarized Madin-Darby canine kidney cells stably expressing MDR1 with ML-7 produced a significant increase in apical to basal permeability and a corresponding decrease in the efflux ratio (threefold; p < 0.01) of [H-3]-digoxin, a classic MDR1 substrate. Together these data show that MDR1-mediated transport of [H-3]-digoxin can be modulated by pharmacological manipulation of myosin RLC, but direct MDR1-RLC interactions are atypical and not explained by the structure of the myosin II holoenzyme. (C) 2010 Elsevier Inc. All rights reserved.