Importin-β negatively regulates multiple aspects of mitosis including RANGAP1 recruitment to kinetochores

Importin-β is the main vector for interphase nuclear protein import and plays roles after nuclear envelope breakdown. Here we show that importin-β regulates multiple aspects of mitosis via distinct domains that interact with different classes of proteins in human cells. The C-terminal region (which binds importin-α) inhibits mitotic spindle pole formation. The central region (harboring nucleoporin-binding sites) regulates microtubule dynamic functions and interaction with kinetochores. Importin-β interacts through this region with NUP358/RANBP2, which in turn binds SUMO-conjugated RANGAP1 in nuclear pores. We show that this interaction continues after nuclear pore disassembly. Overexpression of importin-β, or of the nucleoporin-binding region, inhibited RANGAP1 recruitment to mitotic kinetochores, an event that is known to require microtubule attachment and the exportin CRM1. Co-expressing either importin-β-interacting RANBP2 fragments, or CRM1, restored RANGAP1 to kinetochores and rescued importin-β-dependent mitotic dynamic defects. These results reveal previously unrecognized importin-β functions at kinetochores exerted via RANBP2 and opposed by CRM1.