Cox Charles D, Bae Chilman, Ziegler Lynn, Hartley Silas, Nikolova-Krstevski Vesna, Rohde Paul R, Ng Chai-Ann, Sachs Frederick, Gottlieb Philip A, Martinac Boris
Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales 2010, Australia.
Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.
Nat Commun. 2016 Jan 20;7:10366. doi: 10.1038/ncomms10366.
Mechanosensitive ion channels are force-transducing enzymes that couple mechanical stimuli to ion flux. Understanding the gating mechanism of mechanosensitive channels is challenging because the stimulus seen by the channel reflects forces shared between the membrane, cytoskeleton and extracellular matrix. Here we examine whether the mechanosensitive channel PIEZO1 is activated by force-transmission through the bilayer. To achieve this, we generate HEK293 cell membrane blebs largely free of cytoskeleton. Using the bacterial channel MscL, we calibrate the bilayer tension demonstrating that activation of MscL in blebs is identical to that in reconstituted bilayers. Utilizing a novel PIEZO1-GFP fusion, we then show PIEZO1 is activated by bilayer tension in bleb membranes, gating at lower pressures indicative of removal of the cortical cytoskeleton and the mechanoprotection it provides. Thus, PIEZO1 channels must sense force directly transmitted through the bilayer.
机械敏感离子通道是将机械刺激与离子通量耦合的力传导酶。理解机械敏感通道的门控机制具有挑战性，因为通道所感知的刺激反映了膜、细胞骨架和细胞外基质之间共享的力。在这里，我们研究机械敏感通道PIEZO1是否通过双层膜的力传递而被激活。为了实现这一点，我们生成了基本上不含细胞骨架的HEK293细胞膜泡。使用细菌通道MscL，我们校准了双层膜张力，证明膜泡中MscL的激活与重构双层膜中的激活相同。然后，利用一种新型的PIEZO1-GFP融合蛋白，我们表明PIEZO1在膜泡膜中被双层膜张力激活，在较低压力下开启，这表明皮质细胞骨架及其提供的机械保护被去除。因此，PIEZO1通道必须直接感知通过双层膜传递的力。
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