Yang I C, Cheng T H, Wang F, Price E M, Hwang T C
Department of Physiology, University of Missouri-Columbia 65211, USA.
Am J Physiol. 1997 Jan;272(1 Pt 1):C142-55. doi: 10.1152/ajpcell.1997.272.1.C142.
Modulation of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel by calyculin A and genistein was studied in Hi-5 insect cells infected with baculovirus containing the wild-type CFTR cDNA. In cell-attached patches, CFTR channel activity was not observed until stimulated by forskolin in 90% of the cells, suggesting a low level of basal adenosine 3',5'-cyclic monophosphate activity. Calyculin A, a specific inhibitor of phosphatases 1 and 2A, increased forskolin-induced CFTR activity by 17.2-fold. CFTR channel currents did not deactivate completely after forskolin was withdrawn in the continued presence of calyculin A. Genistein enhanced forskolin-induced CFTR activity by 44.9-fold but could neither activate the CFTR by itself nor prevent complete deactivation on removal of forskolin. Genistein together with calyculin A could adequately prevent deactivation of CFTR currents. Noise analysis of the macroscopic CFTR currents revealed significant differences in the mean current-variance-relationship and the corner frequency of the noise spectra between currents activated by forskolin plus genistein and those activated by forskolin plus calyculin A. Furthermore, genistein enhanced CFTR activity induced by saturating concentrations of forskolin and calyculin A. Our results suggest that genistein and calyculin A modulate the CFTR by different mechanisms and that genistein might inhibit calyculin A-insensitive dephosphorylation of the CFTR.
在感染了含有野生型囊性纤维化跨膜传导调节因子（CFTR）cDNA的杆状病毒的Hi-5昆虫细胞中，研究了毛喉素A和染料木黄酮对CFTR氯离子通道的调节作用。在细胞贴附式膜片钳实验中，直到用福斯可林刺激90%的细胞后才观察到CFTR通道活性，这表明基础腺苷3',5'-环磷酸活性水平较低。毛喉素A是磷酸酶1和2A的特异性抑制剂，可使福斯可林诱导的CFTR活性增加17.2倍。在持续存在毛喉素A的情况下，撤去福斯可林后CFTR通道电流并未完全失活。染料木黄酮可使福斯可林诱导的CFTR活性增加44.9倍，但它自身既不能激活CFTR，也不能在撤去福斯可林时阻止其完全失活。染料木黄酮与毛喉素A共同作用可充分防止CFTR电流失活。对宏观CFTR电流的噪声分析显示，福斯可林加染料木黄酮激活的电流与福斯可林加毛喉素A激活的电流在平均电流-方差关系和噪声谱的转角频率上存在显著差异。此外，染料木黄酮可增强饱和浓度的福斯可林和毛喉素A诱导的CFTR活性。我们的结果表明，染料木黄酮和毛喉素A通过不同机制调节CFTR，并且染料木黄酮可能抑制CFTR的毛喉素A不敏感的去磷酸化作用。
