Yin Wu, Yin Fang-Zhou, Shen Wei-Xing, Cai Bao-Chang, Hua Zi-Chun
The State Key Laboratory of Pharmaceutical Biotechnology, College of Life Science, Nanjing University, Nanjing 210093, PR China.
Int J Biochem Cell Biol. 2008;40(5):942-53. doi: 10.1016/j.biocel.2007.10.036. Epub 2007 Nov 17.
We have previously implicated reactive oxygen species oxygen (ROS) as a critical signal transducer in the upregulation of Na,K-ATPase by low K+ in MDCK cells, but how ROS mediate this process has not been well defined. We reported here that both of hydrogen peroxide (H2O2) and superoxide anion (O2*(-)) were rapidly produced at the early stage of low K+-treated MDCK cells. Further analysis revealed that NADP/NADPH oxidase-derived H2O2 was specifically involved in low K+-induced Na,K-ATPase alpha1 gene transcription as well as alpha1 and beta1 subunits expressions. Exogenous H2O2 even mimicked the stimulatory effect of low K+ on Na,K-ATPase alpha1 gene transcription. Low K+ triggered a H2O2-dependent ERK1/2 phosphorylation in MDCK cells, nonetheless, this ERK1/2 activation did not finally lead to the upregulation of Na,K-ATPase. Similar to previous findings that Na,K-ATPase beta1 gene transcription was mediated by Sp1, Na,K-ATPase alpha1 gene transcription in low K+-treated MDCK cells was also closely relevant to Sp1 participation, as confirmed by siRNA as well as PCR mutagenesis technologies. Furthermore, Sp1 activation was dependent on H2O2 generation triggered by low K+. Taken together, the data described in this study outlines an essential role of H2O2 and Sp1 in mediating the upregulation of Na,K-ATPase in MDCK cells by low external K+.
我们之前已经表明，活性氧（ROS）是MDCK细胞中低钾上调钠钾ATP酶的关键信号转导分子，但ROS如何介导这一过程尚未明确。我们在此报告，在低钾处理的MDCK细胞早期，过氧化氢（H2O2）和超氧阴离子（O2*(-)）均迅速产生。进一步分析表明，NADP/NADPH氧化酶衍生的H2O2特异性参与低钾诱导的钠钾ATP酶α1基因转录以及α1和β1亚基的表达。外源性H2O2甚至模拟了低钾对钠钾ATP酶α1基因转录的刺激作用。低钾在MDCK细胞中触发了H2O2依赖性的ERK1/2磷酸化，然而，这种ERK1/2激活最终并未导致钠钾ATP酶的上调。与之前关于钠钾ATP酶β1基因转录由Sp1介导的发现类似，低钾处理的MDCK细胞中钠钾ATP酶α1基因转录也与Sp1的参与密切相关，这已通过siRNA以及PCR诱变技术得到证实。此外，Sp1的激活依赖于低钾触发的H2O2生成。综上所述，本研究中描述的数据概述了H2O2和Sp1在介导低钾条件下MDCK细胞中钠钾ATP酶上调方面的重要作用。
