Our study demonstrates that during the meiosis to mitosis transition, changes in MTOC organization, spindle assembly, and characteristics and in cell division time do not occur abruptly upon fertilization but progress gradually instead throughout the first eight divisions of the preimplantation mouse embryo. The transition can be subdivided into three phases (): (1) the first three embryonic divisions, when the mechanism of acentrosomal spindle formation is largely shared with meiotic division; (2) the divisions from eight-cell stage until the blastocyst, in which multiple MTOCs or potentially some centrosomes are focused into a sharp bipolar spindle; and (3) the divisions after blastocyst, in which two centrosomes assemble a typical mitotic spindle. The first three divisions in the mouse embryo partially share the mechanism of meiotic spindle formation (summarized in , after NEBD). Given the comparable number and distribution pattern of MTOCs from MII oocyte to zygotes, presumably zygotes inherit MTOCs from oocytes but not from sperm. Nonetheless, the first three divisions also exhibit a change toward mitosis with respect to MTOC maturation and spindle assembly (, before NEBD). In the zygote, MTOCs are recruited for spindle assembly only from the vicinity of the pronuclei, whereas recruitment in the oocyte is from throughout the cytoplasm (Schuh and Ellenberg, 2007
), possibly caused by the lack of the counteracting kinesin-5 force in oocytes. This shift toward mitosis could also account for the lack of microtubule ball formation upon NEBD in zygotes.
Figure 7. Gradual transition from meiotic to mitotic spindle assembly throughout the preimplantation stage in the mouse embryo. Summary of the progressive transition from meiosis to mitosis throughout mouse preimplantation development. See Discussion for details. (more ...)
The second phase, from eight-cell to 64-cell stage, is the time of transition from meiosis-like divisions to mitotic divisions. The number of MTOCs per cell progressively decreases during the preimplantation stage, whereas the total amount of MTOC material per embryo, as estimated by multiplying the mean MTOC number with the volume (), is 3.2, 2.2, 0.5, and 1.8 (arbitrary units) in the zygote, two-cell, eight-cell, and E3.5 stage embryos, respectively, compared with 20.2 in the E4.5 blastocyst. Thus, it is plausible that until blastocyst, noncentriolar MTOCs are generated by splitting the limited amount of materials inherited from the oocyte and available in the embryo in a manner similar to the centrosomal material reported for early Caenorhabditis elegans
embryos (Greenan et al., 2010
; Decker et al., 2011
). When MTOC materials available in a cell become too few and weak, the centrosome generation might be activated. The precise evaluation of this model in the mouse embryo awaits further studies.
The regulation of spindle length according to cell size becomes active as soon as the ratio approaches 1.6, around the fourth division, similar to that observed in HeLa cells (1.4; Goshima and Scholey, 2010
). Although the apparent plateau in spindle length during earlier divisions suggests an upper limit in metaphase spindle length, in agreement with Wühr et al. (2008)
, we cannot exclude the possible scaling correlation between the anaphase spindle length and cell size (Hara and Kimura, 2009
) because of the difficulty in defining the spindle length in anaphase in which the chromosomes move toward the daughter cell poles beyond the eventual nuclear position in the early mouse embryo (e.g., Video 9). Overall, the transition is gradual during this period, not sharp or step wise, and not synchronous in timing, as indicated by the asynchronous emergence of cells with centrin-positive MTOCs (centrosomes) in E3.5 blastocysts.
In the third phase, i.e., after the 64-cell stage, the centriole is present (Gueth-Hallonet et al., 1993
), and the spindle is clearly focused with a well-defined axis at prometaphase. All of these features define cell division later than the blastocyst stage as typical mitosis.
Mouse preimplantation development is not only viewed as a unique phase in terms of developmental mechanisms (O’Farrell et al., 2004
; Motosugi et al., 2005
) but also provides a unique system to study the transition from meiotic to mitotic cell division (this study; Kubiak et al., 2008
; FitzHarris, 2009
). This transition could possibly apply to other organisms: In human zygotes, centrioles are introduced by the sperm (Simerly et al., 1995
) and are detectable by electron microscopy in one of the spindle poles (Sathananthan et al., 1991
). The essential role of the centriole for spindle assembly in early human embryos, however, remains to be shown, as parthenogenetically activated embryos can develop up to the blastocyst stage (Paffoni et al., 2007
; de Fried et al., 2008
Conceivably, in mouse development, the molecular components necessary for reestablishment of the centriole and centrosome are produced during the preimplantation stage, given the lack of evidence for centriole propagation. It will be of particular interest to determine molecules operating during this progressive transition and the stage they become active. The trigger for de novo centriole formation in the blastocyst remains to be investigated but might reflect an exhausted supply of some MTOC components, transcriptional activation of key centriolar components during preimplantation development, or reaching threshold levels in some of the progressively changing cellular features. Because the mouse preimplantation embryo exhibits de novo centrosome formation under physiological conditions, it is an attractive system to investigate the mechanism of centrosome biogenesis and propagation. Future studies promise to identify the components essential for centrosome generation and propagation.