ABSTRACT

The mechanosensitive channel Piezo1 is a crucial membrane mechanosensor ubiquitously expressed in mammalian cell types. Critical to its function in mechanosensory transduction is its ability to change conformation in response to applied mechanical force. Here, we interrogate the role of the anchor domain in the mechanically induced gating of human Piezo1 channels. Using the insertion of glycine residues at each corner of the triangular-shaped anchor domain to decouple this domain we provide evidence that the anchor is important in Piezo1 mechano-gating. Insertion of two extra glycine residues between the anchor and the outer helix of human Piezo1 causes abrogated inactivation and reduced mechanosensitivity. Whereas inserting two glycine residues at the apex of the anchor domain at the conserved amino acid P2113 causes the channel to be more sensitive to membrane forces. Correlation of stretch sensitivity with the volume of the neighboring amino acid, natively a phenylalanine (F2114), suggests this is caused by removal of steric hindrance on the inner pore-lining helix. Smaller volume amino acids at this residue increase sensitivity whereas larger volume reduces mechanosensitivity. The combined data show that the anchor domain is a critical region for Piezo1-mediated force transduction.

 抽象的

机械敏感通道 Piezo1 是一种重要的膜机械传感器，在哺乳动物细胞类型中普遍表达。其在机械感觉转导中的功能的关键是其响应施加的机械力而改变构象的能力。在这里，我们探讨了锚定域在人类 Piezo1 通道机械诱导门控中的作用。通过在三角形锚定结构域的每个角插入甘氨酸残基来解耦该结构域，我们提供了证据表明锚定在 Piezo1 机械门控中很重要。在人 Piezo1 的锚点和外螺旋之间插入两个额外的甘氨酸残基会导致失活消除并降低机械敏感性。而在保守氨基酸 P2113 处的锚定结构域顶端插入两个甘氨酸残基会导致通道对膜力更加敏感。拉伸敏感性与邻近氨基酸（天然苯丙氨酸（F2114））体积的相关性表明，这是由于去除了内孔衬里螺旋上的空间位阻而引起的。该残基上较小体积的氨基酸会增加敏感性，而较大体积的氨基酸会降低机械敏感性。综合数据表明锚定域是 Piezo1 介导的力传导的关键区域。