Investigating the stepping mechanism of myosin X using single molecule tracking and building magnetic tweezers for the measurement of single cell mechanics

Abstract

Myosin X is the first reported myosin to form the anti-parallel dimer in myosin family. It was reported to be localized to the tips of filopodia and to assist in the formation of filopodia. The structure, stepping mechanism and functions of myosin X are crucial for understanding the detailed signal transduction during cell migrations. Here, I focus on the stepping mechanism of myosin X which has been under heated debate. By implementing the single-molecule tracking method called FIONA, the center of the single fluorescence molecule, which used to label myosin X, can be localized with one nanometer accuracy. I found that myosin X has broad distributions of step sizes and frequent backward steps compared with the other members in myosin family, which suggests a unique stepping mechanism on actin bundles along the filopodia. Magnetic tweezers can exert force or torque on biological systems including molecules and cells. It has a wider force range from pico-newton to nano-newton than the other micromanipulation tools such as AFM and optical tweezers. Magnetic tweezers with permanent magnets combined with the fast CMOS camera enable us to investigate the dynamics and mechanics of single DNA whereas the magnetic tweezers with electromagnets allow the investigation of mechanics of single cells. I built a magnetic tweezers with both the permanent magnets and the electromagnet and applied the electromagnetic system into measuring cell mechanical properties. The results of cells measurement are consistent with the previous reports.

Date of Award	2017
Original language	English
Awarding Institution	The Hong Kong University of Science and Technology