The nucleoporin Nup98 is a site for GDP/GTP exchange on Ran and termination of karyopherin β2-mediated nuclear import

Karyopherin beta 2 (Kap beta 2, transportin) binds the MS sequence of human ribonucleoprotein Al and mediates its nuclear import. Here we show a role for the nucleoporin Nup98 in the disassembly of Kap beta 2 import complexes at the nuclear side of the nuclear pore complex (NPC). Kap beta 2 bound to a region at the N terminus of Nup98 that contains an MS-like sequence. The human ribonucleoprotein Al MS sequence competed with Nup98 for binding to Kap beta 2, indicating that Nup98 can dissociate Kappa from its substrate. Binding of Kap beta 2 to Nup98 was inhibited by Ran loaded with guanylyl imidophosphate, suggesting that RanGTP dissociates Kappa from Nup98, RanGTP is produced from RanGDP through nucleotide exchange mediated by RanGEF (RCC1). Immunoelectron microscopy and nucleotide exchange assays revealed functional RanGEF on both sides of the NPC. On the nuclear side, the localization of RanGEF coincided with that of Nup98. RanGEF bound to Nup98 at a region adjacent to the Kappa-binding site. These findings suggest a model where 1) import substrate is released from Kap beta 2 at the nucleoplasmic side of the NPC by competition with the Nup98 MS-like site, 2) Nup98-bound RanGEF catalyzes the formation of RanGTP, and 3) RanGTP dissociates Kappa from Nup98 allowing repeated cycles of import.