Ran Induces Spindle Assembly by Reversing the Inhibitory Effect of Importin α on TPX2 Activity

spindle assembly in Xenopus egg extracts in the pres-European Molecular Biology Laboratory ence of inhibitors of chromosome-induced microtubule Meyerhofstrasse 1 nucleation (Carazo-Salas et al., 2001). These results D-69117 Heidelberg support the hypothesis that the ability of chromosomes Germany to nucleate microtubules is both required and sufficient to form a proper spindle. Although it has been known for some time that chro-Summary mosomes can initiate microtubule assembly and spindle formation (Karsenti et al., 1984a, 1984b) it is only recently The small GTPase Ran, bound to GTP, is required for that insight into the mechanism of this process has been the induction of spindle formation by chromosomes gained. A number of studies showed that the presence in M phase. High concentrations of Ran.GTP are proof a sufficiently high concentration of the small GTPase posed to surround M phase chromatin. We show that Ran in the GTP bound state caused microtubule nucle-the action of Ran.GTP in spindle formation requires ation in Xenopus meiotic cell extract and led to the TPX2, a microtubule-associated protein previously organization of spindle-like structures in the absence of known to target a motor protein, Xklp2, to microtu-either chromosomes or centrosomes (Carazo-Salas et bules. TPX2 is normally inactivated by binding to the Strong evidence that from importin ␣ by the action of Ran.GTP. TPX2 is Ran.GTP also plays a role in spindle formation in vivo in required for Ran.GTP and chromatin-induced microtu-cultured mammalian cells has recently been presented bule assembly in M phase extracts and mediates spon-(Guarguaglini et al., 2000). taneous microtubule assembly when present in ex-The nucleotide (i.e., GTP or GDP) bound state of Ran cess over free importin ␣. Thus, components of the is controlled by the combined action of several proteins. nuclear transport machinery serve to regulate spindle The GTP hydrolysis rate of Ran is greatly stimulated by formation in M phase. its GTPase activating protein, RanGAP (Bischoff et al., 1994). A further stimulation is achieved in the presence Introduction of either RanBP1 or RanBP2, two related Ran.GTP binding proteins (Vetter et al., 1999 and refs. therein). Effi-The transition from interphase to mitosis or meiosis is cient exchange of GDP for GTP on Ran requires the accompanied by a dramatic reorganization of the microtu-guanine nucleotide exchange factor RanGEF or RCC1 bule cytoskeleton. The nucleation of new cytosolic micro-(Bischoff and Ponstingl, 1991; Klebe et al., 1995). RCC1 tubules can no longer occur, leading to the loss of the …