Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba, 260-8675, Japan.
Biochem Pharmacol. 2011 Jan 1;81(1):178-84. doi: 10.1016/j.bcp.2010.09.016. Epub 2010 Sep 22.
Multidrug resistance-associated protein 2 (Mrp2) is an ATP-dependent export pump that mediates the formation of bile-salt-independent bile flow. Disruption of the canalicular localization of Mrp2, without changes in its expression, is observed in chronic liver failure and is accompanied by oxidative stress. We reported previously that Mrp2 is rapidly internalized from the canalicular membrane during acute oxidative stress induced by lipopolysaccharide (LPS) in the rat liver. A disturbance in the colocalization of Mrp2 and radixin (which crosslinks actin with interacting membrane proteins) and endocytic retrieval of Mrp2 are present in chronic liver failure. However, the C-terminal phosphorylation status of radixin (p-radixin; functional form) and its protein-protein interaction with Mrp2 were not examined in the pathological cholestatic situation. In this study, we examined whether the C-terminal phosphorylation status of radixin and its interaction with Mrp2 were affected by LPS-induced experimental liver failure with cholestasis, and whether this condition was accompanied by Mrp2 internalization in the rat liver. At 3h after LPS treatment, the canalicular expression of Mrp2 was decreased, without variation of the other canalicular transporters. Similarly, the canalicular localization of radixin was decreased after LPS treatment. These results show that LPS treatment decreased the total amount of the active form of p-radixin and the amount of radixin that coimmunoprecipitated with Mrp2, and that LPS treatment impaired the protein-protein interaction between Mrp2 and radixin. In conclusion, LPS-induced cholestasis seems to be caused by posttranscriptional regulation of Mrp2, which is due to the disruption of its interaction with radixin and by its dephosphorylation.
多药耐药相关蛋白 2（Mrp2）是一种 ATP 依赖性输出泵，介导胆盐非依赖性胆汁流的形成。在慢性肝功能衰竭中，观察到 Mrp2 的胆小管定位破坏，而其表达没有变化，同时伴有氧化应激。我们之前报道过，在 LPS 诱导的大鼠肝脏急性氧化应激期间，Mrp2 从胆小管膜快速内化。在慢性肝功能衰竭中，存在 Mrp2 和根蛋白（将肌动蛋白与相互作用的膜蛋白交联）的共定位紊乱和 Mrp2 的内吞回收。然而，在病理胆淤积情况下，根蛋白的 C 端磷酸化状态（功能性形式）及其与 Mrp2 的蛋白-蛋白相互作用尚未被检查。在这项研究中，我们检查了 LPS 诱导的实验性肝衰竭伴胆汁淤积是否会影响根蛋白的 C 端磷酸化状态及其与 Mrp2 的相互作用，以及这种情况是否伴有大鼠肝脏中 Mrp2 的内化。在 LPS 处理后 3 小时，Mrp2 的胆小管表达减少，而其他胆小管转运体没有变化。同样，根蛋白的胆小管定位在 LPS 处理后减少。这些结果表明，LPS 处理降低了 p-radixin 的活性形式的总量和与 Mrp2 共免疫沉淀的根蛋白的量，并且 LPS 处理破坏了 Mrp2 和根蛋白之间的蛋白-蛋白相互作用。总之，LPS 诱导的胆汁淤积似乎是由于 Mrp2 的转录后调节引起的，这是由于其与根蛋白相互作用的破坏和去磷酸化。
