Force Generation by Dynamic Microtubules In Vitro

Biopolymers are essential for cellular organization. They bridge the cell interior, forming a framework that is used as a reference for different cellular organelles. This framework, called the cytoskeleton, is not static but constantly reorganizes. The dynamics of the cytoskeleton allows the cell to rearrange its interior for various processes, such as cell division. This dynamic reorganization relies at least partly on forces that arise from the assembly and disassembly of cytoskeletal biopolymers. In many cases, these forces are generated when biopolymers interact with the cell boundary. This chapter focuses on force generation by and regulation of microtubules (MTs) that interact with growth-opposing barriers. We describe three in vitro assays that can be used to mimic MT interactions with the cell boundary. The essential components in each of our minimal systems are (functionalized) microfabricated barriers against which we grow MTs under different conditions. We describe in detail the different methods and assays necessary to realize these in vitro experiments.