Dynamic Actin Networks Utilized to Support Simultaneous Processive Motility of Two Different Myosin Motor Classes

Abstract

In an eukaryotic cell multiple myosin motor families coexist and use the same actin cytoskeleton as tracks for processive motility and transport of cargoes. It is unclear how these motors interact while sharing the same F-actin and whether there are exclusion mechanisms that prioritize function of one myosin class over the other. To assess the influence that different myosin classes have on each other, we developed an assay that combines two dynamic systems: elongating actin filaments with distinguished barbed and pointed ends, and myosins moving along these filaments.In this study two different myosins motors were used: myosin-5 and myosin-6. These two motors have distinct functions in the cell and are known to travel in opposite directions along actin filaments. Myosin-5 walks towards the barbed end of F-actin towards dynamically rearranging actin in the cell periphery. Myosin-6, which is used in endocytic traffic, is a pointed-end motor that walks towards the cell center. We successfully reconstituted simultaneous bidirectional motility of myosin-5 and myosin-6 on single polymerizing filaments of actin. Speed, run lengths, and effective landing rates of the motors in this ‘two motor assay’ were compared to ‘single motor assays’ featuring motility of myosin-5 or myosin-6 exclusively. We also report the frequency and outcomes of collision events between motors walking in opposite directions along single actin filament.