Reconstitution of three-phase microtubule polymerisation dynamics

Cytoplasmic microtubules (MTs) undergo growth, shrinkage, and pausing. However, how MT polymerisation cycles are produced and spatiotemporally regulated at a molecular level is unclear, as the entire cycle has not been recapitulated in vitro with defined components. In this study, we reconstituted dynamic MT plus end behaviour involving all three phases, by mixing tubulin with five Drosophila proteins, EB1, XMAP215Msps, Sentin, kinesin-13Klp10A, and CLASPMast/Orbit. When singly mixed with tubulin, CLASPMast/Orbit strongly inhibited MT catastrophe and reduced the growth rate. However, in the presence of the other four factors, CLASPMast/Orbit acted as an inducer of pausing. The mitotic kinase