Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
???displayArticle.abstract???
Female meiotic spindles in many organisms form in the absence of centrosomes, the organelle typically associated with microtubule (MT) nucleation. Previous studies have proposed that these meiotic spindles arise from RanGTP-mediated MT nucleation in the vicinity of chromatin; however, whether this process is sufficient for spindle formation is unknown. Here, we investigated whether a recently proposed spindle-based MT nucleation pathway that involves augmin, an 8-subunit protein complex, also contributes to spindle morphogenesis. We used an assay system in which hundreds of meiotic spindles can be observed forming around chromatin-coated beads after introduction of Xenopus egg extracts. Spindles forming in augmin-depleted extracts showed reduced rates of MT formation and were predominantly multipolar, revealing a function of augmin in stabilizing the bipolar shape of the acentrosomal meiotic spindle. Our studies also have uncovered an apparent augmin-independent MT nucleation process from acentrosomal poles, which becomes increasingly active over time and appears to partially rescue the spindle defects that arise from augmin depletion. Our studies reveal that spatially and temporally distinct MT generation pathways from chromatin, spindle MTs, and acentrosomal poles all contribute to robust bipolar spindle formation in meiotic extracts.
Fig. 4. Visualization of growing MT plus ends via GFP-EB1 in extracts with control antibody (Left) or anti-Dgt4 antibody (Right). (A) Representative images of tubulin (Cy3-channel) and GFP–EB1 (140 nM) are shown for extract incubated with control or Dgt4 antibody (see Fig. 3G and Fig. S3 C–E) at three assembly stages (see Fig. 1D). (B and C) Augmin-inhibited spindles show a reduction of EB1–GFP comets from the middle of the spindles. Note the prominent density of EB1–GFP comets at the acentrosomal poles in C (both control and Dgt4 AB). (Scale bars, 10 μm.) See also Movie S3 (control AB) and Movie S4 (Dgt4 AB). (D) Quantitation of the ratio between the EB1 intensity in the midzone and near the poles (see circles for zone of measurement). Mean and SEM are derived from three independent experiments (2–7 spindles measured for each time point in each independent experiment).
Albee,
Xenopus TACC3/maskin is not required for microtubule stability but is required for anchoring microtubules at the centrosome.
2008, Pubmed,
Xenbase
Albee,
Xenopus TACC3/maskin is not required for microtubule stability but is required for anchoring microtubules at the centrosome.
2008,
Pubmed
,
Xenbase
Carazo-Salas,
Generation of GTP-bound Ran by RCC1 is required for chromatin-induced mitotic spindle formation.
1999,
Pubmed
,
Xenbase
Clausen,
Self-organization of anastral spindles by synergy of dynamic instability, autocatalytic microtubule production, and a spatial signaling gradient.
2007,
Pubmed
,
Xenbase
Dinarina,
Chromatin shapes the mitotic spindle.
2009,
Pubmed
,
Xenbase
Goshima,
Augmin: a protein complex required for centrosome-independent microtubule generation within the spindle.
2008,
Pubmed
Goshima,
Genes required for mitotic spindle assembly in Drosophila S2 cells.
2007,
Pubmed
Groen,
Functional overlap of microtubule assembly factors in chromatin-promoted spindle assembly.
2009,
Pubmed
,
Xenbase
Groen,
XRHAMM functions in ran-dependent microtubule nucleation and pole formation during anastral spindle assembly.
2004,
Pubmed
,
Xenbase
Hannak,
Investigating mitotic spindle assembly and function in vitro using Xenopus laevis egg extracts.
2006,
Pubmed
,
Xenbase
Heald,
Self-organization of microtubules into bipolar spindles around artificial chromosomes in Xenopus egg extracts.
1996,
Pubmed
,
Xenbase
Heald,
Spindle assembly in Xenopus egg extracts: respective roles of centrosomes and microtubule self-organization.
1997,
Pubmed
,
Xenbase
Houghtaling,
Op18 reveals the contribution of nonkinetochore microtubules to the dynamic organization of the vertebrate meiotic spindle.
2009,
Pubmed
,
Xenbase
Khodjakov,
Centrosome-independent mitotic spindle formation in vertebrates.
2000,
Pubmed
Kollman,
Microtubule nucleating gamma-TuSC assembles structures with 13-fold microtubule-like symmetry.
2010,
Pubmed
Lawo,
HAUS, the 8-subunit human Augmin complex, regulates centrosome and spindle integrity.
2009,
Pubmed
Loughlin,
A computational model predicts Xenopus meiotic spindle organization.
2010,
Pubmed
,
Xenbase
Mahoney,
Making microtubules and mitotic spindles in cells without functional centrosomes.
2006,
Pubmed
Maiato,
Kinetochore-microtubule interactions during cell division.
2004,
Pubmed
Megraw,
Zygotic development without functional mitotic centrosomes.
2001,
Pubmed
Merdes,
A complex of NuMA and cytoplasmic dynein is essential for mitotic spindle assembly.
1996,
Pubmed
,
Xenbase
Mitchison,
Dynamic instability of microtubule growth.
,
Pubmed
Mitchison,
Bipolarization and poleward flux correlate during Xenopus extract spindle assembly.
2004,
Pubmed
,
Xenbase
Murray,
Cyclin synthesis drives the early embryonic cell cycle.
1989,
Pubmed
,
Xenbase
Samejima,
Two distinct regions of Mto1 are required for normal microtubule nucleation and efficient association with the gamma-tubulin complex in vivo.
2008,
Pubmed
Uehara,
The augmin complex plays a critical role in spindle microtubule generation for mitotic progression and cytokinesis in human cells.
2009,
Pubmed
Uehara,
Functional central spindle assembly requires de novo microtubule generation in the interchromosomal region during anaphase.
2010,
Pubmed
Verde,
Taxol-induced microtubule asters in mitotic extracts of Xenopus eggs: requirement for phosphorylated factors and cytoplasmic dynein.
1991,
Pubmed
,
Xenbase
Walczak,
Mechanisms of mitotic spindle assembly and function.
2008,
Pubmed
,
Xenbase
Wiese,
Microtubule nucleation: gamma-tubulin and beyond.
2006,
Pubmed
Wilde,
Stimulation of microtubule aster formation and spindle assembly by the small GTPase Ran.
1999,
Pubmed
,
Xenbase
