Berdiev B K, Prat A G, Cantiello H F, Ausiello D A, Fuller C M, Jovov B, Benos D J, Ismailov I I
Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, Alabama 35294-0005, USA.
J Biol Chem. 1996 Jul 26;271(30):17704-10. doi: 10.1074/jbc.271.30.17704.
Cytoskeletal elements play an important role in the regulation of ion transport in epithelia. We have studied the effects of actin filaments of different length on the alpha, beta, gamma-rENaC (rat epithelial Na+ channel) in planar lipid bilayers. We found the following. 1) Short actin filaments caused a 2-fold decrease in unitary conductance and a 2-fold increase in open probability (Po) of alpha,beta,gamma-rENaC. 2) alpha,beta,gamma-rENaC could be transiently activated by protein kinase A (PKA) plus ATP in the presence, but not in the absence, of actin. 3) ATP in the presence of actin was also able to induce a transitory activation of alpha, beta,gamma-rENaC, although with a shortened time course and with a lower magnitude of change in Po. 4) DNase I, an agent known to prohibit elongation of actin filaments, prevented activation of alpha,beta,gamma-rENaC by ATP or PKA plus ATP. 5) Cytochalasin D, added after rundown of alpha,beta,gamma-rENaC activity following ATP or PKA plus ATP treatment, produced a second transient activation of alpha,beta,gamma-rENaC. 6) Gelsolin, a protein that stabilizes polymerization of actin filaments at certain lengths, evoked a sustained activation of alpha,beta,gamma-rENaC at actin/gelsolin ratios of <32:1, with a maximal effect at an actin/gelsolin ratio of 2:1. These results suggest that short actin filaments activate alpha, beta,gamma-rENaC. PKA-mediated phosphorylation augments activation of this channel by decreasing the rate of elongation of actin filaments. These results are consistent with the hypothesis that cloned alpha,beta,gamma-rENaCs form a core conduction unit of epithelial Na+ channels and that interaction of these channels with other associated proteins, such as short actin filaments, confers regulation to channel activity.
细胞骨架成分在上皮细胞离子转运的调节中发挥重要作用。我们研究了不同长度的肌动蛋白丝对平面脂质双分子层中α、β、γ-rENaC（大鼠上皮钠通道）的影响。我们发现如下情况。1）短肌动蛋白丝使α、β、γ-rENaC的单位电导降低2倍，开放概率（Po）增加2倍。2）在有肌动蛋白存在但无肌动蛋白不存在的情况下，蛋白激酶A（PKA）加ATP可短暂激活α、β、γ-rENaC。3）有肌动蛋白存在时，ATP也能够诱导α、β、γ-rENaC的短暂激活，尽管时间进程缩短且Po的变化幅度较小。4）DNase I是一种已知可阻止肌动蛋白丝伸长的试剂，可阻止ATP或PKA加ATP对α、β、γ-rENaC的激活。5）在ATP或PKA加ATP处理后α、β、γ-rENaC活性降低后加入细胞松弛素D，可使α、β、γ-rENaC再次短暂激活。6）凝溶胶蛋白是一种在特定长度稳定肌动蛋白丝聚合的蛋白质，在肌动蛋白/凝溶胶蛋白比例<32:1时可诱发α、β、γ-rENaC的持续激活，在肌动蛋白/凝溶胶蛋白比例为2:1时效果最佳。这些结果表明短肌动蛋白丝激活α、β、γ-rENaC。PKA介导的磷酸化通过降低肌动蛋白丝的伸长速率增强该通道的激活。这些结果与以下假设一致，即克隆的α、β、γ-rENaC形成上皮钠通道的核心传导单位，并且这些通道与其他相关蛋白（如短肌动蛋白丝）的相互作用赋予通道活性调节作用。
