Chromosomal association of Ran during meiotic and mitotic divisions.

Recent studies in Xenopus egg extracts indicate that the small G protein Ran has a central role in spindle assembly and nuclear envelope reformation. We determined Ran localization and dynamics in cells during M phase. By immunofluorescence, Ran is accumulated on the chromosomes of meiosis-II-arrested Xenopus eggs. In living cells, fluorescently labeled Ran associated with the chromosomes in Xenopus and remained associated during anaphase when eggs were artificially activated. Fluorescent Ran associated with chromosomes in mouse eggs, during meiotic maturation and early embryonic divisions in starfish, and to a lesser degree during mitosis of a cultured mammalian cell line. Chromosomal Ran undergoes constant flux. From photobleach experiments in immature starfish oocytes, chromosomal Ran has a k(off) of approximately 0.06 second(-1), and binding analysis suggests that there is a single major site. The chromosomal interactions may serve to keep Ran-GTP in the vicinity of the chromosomes for spindle assembly and nuclear envelope reformation.

A novel centrosome-associated protein with affinity for microtubules.

We have identified a novel mammalian protein, MIR1, with microtubule-binding activity. MIR1 is a relative of MID1/midin, the protein implicated in Opitz G/BBB syndrome. In tissue culture cells, MIR1 is enriched at the centrosome. MIR1 dissociates from centrosomes at the G2/M transition and is recruited back to spindle poles during anaphase. When overexpressed during interphase, MIR1 binds along microtubule filaments, which become stabilized, bundled and detached from the centrosome. In mitosis, overexpressed MIR1 dissociates from microtubules but still affects the normally focused localization of gamma-tubulin in spindle poles. Tight binding to microtubules in interphase appears to require an oligomeric state of MIR1, and phosphorylation in mitosis at predicted cyclin-dependent kinase (cdk) sites weakens the interaction.