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要旨:
In order to segregate chromatin during cell division to the two daughter cells, a bipolar spindle needs to form. The mitotic spindle consists of anti‐parallel microtubule bundles, connecting the microtubule organizing center (MTOC) via the minus‐ends of the microtubules with the kinetochores via the plus ends. The most prominent MTOCs are centrosomes serving as spindle poles during mitosis in vertebrate somatic cells. The centrosomes contain the γ‐tubulin ring complex (γ ‐TURC) which promotes the nucleation of microtubules. Microtubules are highly dynamic, changing the state of polymerization and depolymerization at the plus end. Beside the centrosome nucleation pathway, a chromatin‐ and Ran‐ dependent nucleation pathway is known. The latter functions by a gradient of Ran in its GTP‐bound form ‐ accumulating close to chromatin ‐ to its GDP‐bound version. Ran‐GTP induces microtubule polymerization by releasing microtubule associated proteins from importins locally around chromatin. Mitotic spindle formation can be reconstituted in vitro using Xenopus laevis egg extract. Labeled tubulin incubated in egg extracts polymerizes for example in the presence of sperm chromatin or the dominant positive mutant of Ran. Using this cell‐free system, I found that additional GTPases are involved in microtubule functions. The Developmentally Regulated GTP binding proteins 1 and 2 (DRG1 and 2) as well as their stabilizing proteins DRG family regulatory proteins (DFRP1 and 2) localize to microtubule nucleation centers in egg extracts. Additionally, DRG1 and DFRP1 bind microtubules in vitro. Furthermore, results obtained by addition of eGFP‐tagged DRG1 and DRG2 to the Ran aster assay suggest that these GTPases increase the microtubule polymerization. Although the specific function is not known yet, these results suggest that DRGs play a role in the assembly of the mitotic spindle – most likely by increasing the polymerization.
