Running on Ran Nuclear Transport and the Mitotic Spindle

sol and GTP hydrolysis. In particular, nuclear import Far from being passive participants in the process of is accomplished for proteins bearing classical nuclear mitosis, chromosomes have an active and essential role localization signals (NLS) by a heterodimeric receptor in the assembly of spindles required for their accurate comprised of a Ran-GTP binding protein, importin b, division into the two daughter cells. One way in which and an adaptor protein, importin a (Figure 1A). Importin they exert this influence is through the selective stabilia recognizes NLS-bearing proteins. Importin b binds to zation of microtubules (MTs) in their immediate vicinity, the importin a-NLS complex and facilitates its movethereby contributing to the nucleation and growth of ment through the nuclear pore. Importin b’s use of adapspindle MTs, as well as to their directed elongation from tor subunits in this manner is unusual, since transport nucleating structures such as centrosomes (reviewed receptors generally bind to their cargo directly. Importin in Andersen, 1999). The mechanistic details of how chrob is responsible for the translocation of other cargoes mosomes promote MT stability remained an intriguing through direct association. Importin b dissociates from mystery until the surprising discovery that the Ran importin a-NLS after binding Ran-GTP in the nucleus, GTPase is an essential component of this process (reand returns to the cytosol in association with Ran-GTP. viewed by Kahana and Cleveland, 1999). Importin a dissociates from NLS within the nucleus, and Ran is a very abundant and highly conserved GTPase is then exported to the cytosol in association with an (reviewed in Sazer and Dasso, 2000). Previous data had importin b–related export receptor, CAS, and Ran-GTP. implicated Ran in many aspects of nuclear function, The Relationship between Ran and Mitotic including nuclear transport, cell cycle control, and postSpindle Assembly mitotic nuclear assembly. The finding that Ran regulates MTs are noncovalent polymers, composed of aand spindle assembly immediately posed the question as to b-tubulin dimeric subunits (reviewed in Desai and Mithow a single protein could be involved in such a diverse chison, 1997). MTs alternate abruptly and stochastically array of cellular activities and whether the effectors for between phases of polymerization and depolymerization. each of these roles would be similar or distinct. Recent This behavior has been termed “dynamic instability.” reports published in January issues of Cell (Gruss et al., Dynamic instability is characterized by four parameters: 2001; Nachury et al., 2001) and Science (Wiese et al., polymerization rate, depolymerization rate, frequency of 2001) shed light on this question and allow the proposal transition from growth to shrinkage (catastrophe freof a much more unified view of Ran’s biochemistry quency), and frequency of transition from shrinkage to throughout the cell cycle. growth (rescue frequency). MT dynamics are highly regThe Biochemistry of Ran and Its Role ulated during the cell cycle. The rate of catastrophe in Nuclear Transport increases 5to 10-fold upon entry into mitosis, causing The core biochemistry of Ran is similar to that of many mitotic MTs to be much shorter and more dynamic than Ras-related GTPases (reviewed in Gorlich and Kutay, interphase MTs (Desai and Mitchison, 1997). The MT 1999; Sazer and Dasso, 2000). Ran’s intrinsic rates of nucleation capacity of centrosomes is also markedly nucleotide exchange and hydrolysis are slow. In vivo, enhanced as cells enter mitosis (Kuriyama and Borisy, these reactions require a nucleotide exchange factor 1981). Mitotic chromosomes locally decrease catastrophe (RCC1) and a GTPase activating protein (RanGAP1) to frequency and increase rescue frequency, promoting the occur at physiological rates. A protein called RanBP1 elongation of spindle MTs (Dogterom et al., 1996). A numbinds to Ran-GTP with high affinity, and acts as an ber of experiments have suggested that chromosomeessential accessory factor to increase RanGAP1-mediinduced MT stabilization requires the product of a chromaated nucleotide hydrolysis. During interphase, RCC1 is tin-associated enzyme, which becomes distributed by a chromatin-associated nuclear protein, while the bulk diffusion into a gradient that is inversely proportional to of RanBP1 and RanGAP1 are cytosolic. The asymmetric distance from the chromosome (Andersen, 1999). distribution of nucleotide exchange and hydrolysis enRan-GTP promotes MT polymerization within M phase– zymes across the nuclear envelope predicts that Ranarrested Xenopus egg extracts (Kahana and Cleveland, GTP should be largely nuclear and Ran-GDP should be 1999). The addition of high levels of Ran-GTP, constitulargely cytosolic. This distribution plays a key role in tively GTP-bound Ran mutants (RanQ69L, RanL43E), determining the directionality of nuclear transport. or exogenous RCC1 promotes massive spontaneous The requirement for Ran in nuclear transport has been polymerization of tubulin, even in the absence of sperm extensively studied (reviewed in Gorlich and Kutay, chromatin or centrioles. Ran-GTP-induced MTs become