Fig. 5: Backward transition from anaphase to metaphase-like microtubule bundle organization.

A TIRF microscopy images showing microtubule bundles pre-organized for ~10−15 min in the presence of 20 nM unphosphorylated PRC1-Alexa546 (green), 10 nM KIF4A-mGFP (blue) and 18 µM Alexa647-tubulin (red), followed by addition of 100 nM CDK1/cyclin B/CKS1 to the top reservoir for delivery to the imaging chamber to phosphorylate PRC1. A large fraction of PRC1 and KIF4A was lost within minutes from the compacted antiparallel overlaps. Times in min:sec, scale bars as indicated. B Exemplary snap shot depict the temporal progression of simulated antiparallel bundles of 4 microtubules connected by PRC1 (green) and KIF4A (blue), undergoing a transition from anaphase-to-metaphase-like organization. Microtubule orientation indicated by color (plus ends up: cyan, plus ends down: red). The unbinding rate of PRC1 (k_off) and the microtubule plus end growth speed (v_g) were changed as indicated to trigger reorganization. C Average antiparallel overlap length of 10 experimental (top) and 10 simulated (bottom) microtubule bundles as a function of time. The gray dotted lines indicate the time of addition of CDK1/cyclin B/CKS1 in the experiment, or change of unbinding rate (k_off) and microtubule growth speed in the simulation, as indicated. n = 2 independent experiments, total of 10 antiparallel overlaps (top), and n = 10 independent simulations, total of 10 antiparallel overlaps (bottom). Black and gray lines are mean and individual overlap length, respectively. Source data are provided as a Source Data file.