Mechanosensitivity Of PIEZO Channel Protein In Human Breast Cancer Cell Probed By Atomic Force Microscope

Mechanotransduction,which transforms the mechanical stimulation into biological signals,plays pivotal roles in physiological functions.The recent discovery found PIEZO channel as mechanosensitivity cation channel,which divides into the structure-similar PIEZO1 and PIEZO2 in vertebrate subphylum.Those two proteins only works when they cooperate.PIEZO can sense mechanical stimulation by peripheral blades and transport cations by opening the cap and transport alkali ions such as K+,Na+,Ca2+,Mg2+ and Mn2+.In this study,we found that MDA-MB-231 expresses the mechanosensitive behavior of PIEZO in nature without any extra treatment.In this study,atomic force microscopy has been applied to test biomechanics of PIEZO channel protein in MD A-MB-231 cells.The main works and the conclusions are as follows:(1)The atomic force microscopy cantilever is used to express PIEZO protein with a instant force stimulation,which suggests that PIEZO can express naturally on MDA-MB-231.The concentration of Ca2+ in cell will rise in a very short time,and will deliver this change to neighbouring cells rapidly.(2)The PIEZO mechanical experiment confirms its activation percentage rises with pressing intensity in 100-500nN,and the PIEZO’s expressing intensity is connected with the cytoactive of cell it belongs to.The AFM cantilever vibration experiment confirms that the cytoactive of MD A-MB-231 is changing with experiment time because of the influence of cell cycle.(3)The PIEZO protein is treated with various blockers including GsMTx4,SKF96365,EGTA,and ruthenium red.The result suggests that PIEZO can be specially blocked by GsMTx4,and the experiment of EGTA ruthenium red shows that calcium cation flows from extracellular domain into intracellular domain when PIEZO is activated,which confirms that PIEZO is stretch-activated cation channels.(4)We have also done the AFM scanning and confocal observation of MD A-MB-231 cell membrane,and find that the Young’s module of cell surface changes significanly while its topography maintaines smoothness and its non-core area exists filamentous actin structure,which confirms that cell membrane is supported by cytoskeleteon and mechanism linkage maybe exist between the attached PIEZO protein and cytoskeleteon.The results on PIEZO of MDA-MB-231 can help establish a basis of preventing and controlling of human breast cancer cell via mechanical forces.