Gopalakrishnan S, Raman N, Atkinson S J, Marrs J A
Department of Medicine, Division of Nephrology, Indiana University Medical Center, Indianapolis, Indiana 46202-5116, USA.
Am J Physiol. 1998 Sep;275(3):C798-809. doi: 10.1152/ajpcell.1998.275.3.C798.
Tight junctions control paracellular permeability and cell polarity. Rho GTPase regulates tight junction assembly, and ATP depletion of Madin-Darby canine kidney (MDCK) cells (an in vitro model of renal ischemia) disrupts tight junctions. The relationship between Rho GTPase signaling and ATP depletion was examined. Rho inhibition resulted in decreased localization of zonula occludens-1 (ZO-1) and occludin at cell junctions; conversely, constitutive Rho signaling caused an accumulation of ZO-1 and occludin at cell junctions. Inhibiting Rho before ATP depletion resulted in more extensive loss of junctional components between transfected cells than control junctions, whereas cells expressing activated Rho better maintained junctions during ATP depletion than control cells. ATP depletion and Rho signaling altered phosphorylation signaling mechanisms. ZO-1 and occludin exhibited rapid decreases in phosphoamino acid content following ATP depletion, which was restored on recovery. Expression of Rho mutant proteins in MDCK cells also altered levels of occludin serine/threonine phosphorylation, indicating that occludin is a target for Rho signaling. We conclude that Rho GTPase signaling induces posttranslational effects on tight junction components. Our data also demonstrate that activating Rho signaling protects tight junctions from damage during ATP depletion.
紧密连接控制细胞旁通透性和细胞极性。Rho GTP酶调节紧密连接的组装，而Madin-Darby犬肾（MDCK）细胞（肾缺血的体外模型）的ATP耗竭会破坏紧密连接。研究了Rho GTP酶信号传导与ATP耗竭之间的关系。抑制Rho会导致闭合蛋白-1（ZO-1）和闭合蛋白在细胞连接处的定位减少；相反，组成型Rho信号传导会导致ZO-1和闭合蛋白在细胞连接处积累。在ATP耗竭前抑制Rho导致转染细胞之间的连接成分比对照连接处损失更广泛，而表达活化Rho的细胞在ATP耗竭期间比对照细胞更好地维持连接处。ATP耗竭和Rho信号传导改变了磷酸化信号传导机制。ATP耗竭后，ZO-1和闭合蛋白的磷酸氨基酸含量迅速下降，恢复时恢复。在MDCK细胞中表达Rho突变蛋白也改变了闭合蛋白丝氨酸/苏氨酸磷酸化水平，表明闭合蛋白是Rho信号传导的靶点。我们得出结论，Rho GTP酶信号传导诱导对紧密连接成分的翻译后效应。我们的数据还表明，激活Rho信号传导可保护紧密连接在ATP耗竭期间免受损伤。
