Probing mechanisms for force-sensing in mechanosensitive ion channels with artificial droplet bilayer systems

Jaggers O1, Ridone P2, Xiao B3, Martinac B2 and Baker MAB1

  1. School of Biotechnology and Biomolecular Science, UNSW.
  2. Victor Chang Cardiac Research Institute.
  3. Tsinghua University.

Droplet Hydrogel Bilayers enable simultaneous single channel current and fluorescence measurements[1]. They have been used to characterise the functionality of alpha-haemolysin for use in nucleobase recognition in DNA sequencing and they have been arranged in multiple arrays to parallelise high throughput channel measurements. We recently used this platform to apply force and measure the response of bacterial mechanosensitive ion channel MscL[2]. We are now using this platform to investigate the force sensitive ion channel PIEZO1[3], in which single point mutations cause blood disorders such as xerocytosis and which is generally linked to cancer progression and post traumatic osteoarthritis. We observe that PIEZO1 in DHBs displays the same gating activity in DHBs as in patch-liposomes, and, similarly, is much more sensitive in the presence of cholesterol. We also investigate the use of DNA-origami to regulate lipid-lipid interactions. This serves to enable light-triggered insertion of membrane proteins into artificial bilayers and allows greater spatiotemporal control. [1.] A. J. Heron, et al., JACS. 131, 1652-1653 (2009). [2.] K. Rosholm et al., Scientific Reports. 7, 45180 (2017). [3.] Q Zhao et al., Nature, 554, 487-492 (2018).