Spindle assembly in the absence of chromosomes in mouse oocytes

State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Chaoyang, Beijing 100101, People's Republic of China.
Reproduction (Cambridge, England) (Impact Factor: 3.17). 01/2008; 134(6):731-8. DOI: 10.1530/REP-07-0149
Source: PubMed


This study was carried out to investigate the contributions of chromosomes to spindle assembly in mouse oocytes. We generated two groups of cytoplasts (holo- and hemi-cytoplasts) by enucleation of germinal vesicle (GV), metaphase I (MI), and metaphase II (MII) oocytes using micromanipulation technology. After in vitro culture for 18 h, spindles with different shapes (bi-, mono-, or multipolar) formed in most of these cytoplasts except in hemi-GV cytoplasts. Two or more spindles were observed in most of holo-GV, holo-MI, and holo-MII cytoplasts (76.1, 77.0, and 83.7% respectively). However, the proportions of hemi-MI and hemi-MII cytoplasts with multiple sets of spindles decreased to 17.6 and 20.7% respectively. A single bipolar spindle was observed in each sham-operated oocyte generated by removing different volumes of cytoplasm from the oocytes and keeping nuclei intact. Localization of gamma-tubulin showed that microtubule organizing centers (MTOCs) were dispersed at each pole of the multiple sets of spindles formed in holo-cytoplasts. However, most of the MTOCs aggregated at the two poles of the bipolar spindle in sham-operated oocytes. Our results demonstrate that chromosomes are not essential for initiating spindle assembly but for directing distinct MTOCs to aggregate to form a bipolar spindle. Some factors of undetermined nature may pre-exist in an inactive form in GV-stage ooplasm, serving as initiators of spindle assembly upon their activation. Moreover, GV materials released into the cytoplasm may facilitate spindle assembly in normal meiotic maturation.

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Available from: Yi-liang Miao
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    • "These results demonstrated that meiotic spindle assembly in mouse oocytes is driven by selforganization of MT arrays into bipolar structures (Brunet et al. 1999), similar to spindle assembly in other systems (Karsenti and Vernos 2001). Because the enucleation or bisection of oocytes (Brunet et al. 1999; Yang et al. 2007) could not remove all Ran and RCC1, RanGTP likely supported the observed MT selforganization . It is possible that X. laevis egg extracts are depleted of components responsible for spontaneous MT formation. "
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    • "The formation of a stable bipolar meiotic spindle is crucial for chromosome disjunction in which chromosome attachment, alignment and pulling toward spindle poles are critical steps for accurate segregation. Although chromatin interaction with microtubules appears to influence microtubule dynamics, a bipolar spindle can still form in the absence of chromatin, as is the case in enucleated mouse oocytes [45], [46]. Our studies for the first time showed that phosphorylated MAPKAPK 2 (p-MK2) is localized to bipolar spindle minus ends and chromosomes during metaphase I and metaphase II in meiosis. "
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