Highly Asynchronous and Asymmetric Cleavage Divisions Accompany Early Transcriptional Activity in Pre-Blastula Medaka Embryos Michael Kraeussling . , Toni Ulrich Wagner*., Manfred Schartl Abstract In the initial phase of development of fish embryos, a prominent and critical event is the midblastula transition (MBT). Before MBT cell cycle is rapid, highly synchronous and zygotic gene transcription is turned off. Only during MBT the cell cycle desynchronizes and transcription is activated. Multiple mechanisms, primarily the nucleocytoplasmic ratio, are supposed to control MBT activation. Unexpectedly, we find in the small teleost fish medaka (Oryzias latipes) that at very early stages, well before midblastula, cell division becomes asynchronous and cell volumes diverge. Furthermore, zygotic transcription is extensively activated already after the 64-cell stage. Thus, at least in medaka, the transition from maternal to zygotic transcription is uncoupled from the midblastula stage and not solely controlled by the nucleocytoplasmic ratio. Citation: Kraeussling M, Wagner TU, Schartl M (2011) Highly Asynchronous and Asymmetric Cleavage Divisions Accompany Early Transcriptional Activity in Pre- Blastula Medaka Embryos. PLoS ONE 6(7): e21741. doi:10.1371/journal.pone.0021741 Editor: Carl-Philipp Heisenberg, Institute of Science and Technology Austria, Austria Received November 29, 2010; Accepted June 10, 2011; Published July 7, 2011 Copyright: 2011 Kraeussling et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was supported by German Research Foundation (DFG) via the ‘‘pecial research group 1048’’(http://www.gk-1048.uni-wuerzburg.de/) and by the EU through Plurigenes (FP6 project 018673)(http://www.plurigenes.org/). This publication was funded by the German Research Foundation (DFG) and the University of Wuerzburg in the funding program Open Access Publishing. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: toni.wagner@biozentrum.uni-wuerzburg.de . These authors contributed equally to this work. Introduction The early development of many animals apparently follows a common scheme: it is characterized by a rapid and synchronous sequence of cell divisions [1][2]. During this period, the cleavage- phase, the single cell of the fertilized egg is divided into a large number of blastomers. This bulk of cells will finally form the blastula. While the embryo reaches the blastula stage, the cells pass through an important time point of early development, which is called midblastula transition (MBT). This process changes both characteristics and behavior of the cells. Before MBT, the cell cycle is highly synchronous. It is rapid because the cells lack both G1- and G2-phases [3] and there is no zygotic gene expression [1][4][5][6]. The cell cycle elongates and desynchronizes only after the cells have passed through the MBT. Cells gain motility and start to express the zygotic genome while maternal mRNAs are degraded [7]. The time points of activation of the midblastula transition vary between different species. MBT starts early in Drosophila at stage 4 (embryonic division cycle 11) [8], in Xenopus at stage 8 (cycle 12) [1][3][9][10] , in Zebrafish at stage 10 (cycle 10) [2] and earlier work identified the beginning of MBT for medaka fish at around stage 11 (cycle 11–12) [6]. At present, four models exist that try to explain MBT regulation: the maternal clock [11][12], transcriptional abortion [5][13], chromatin regulation [14][15], and the nucleo-cytoplasmic ratio. Among these, the regulation by a nucleo-cytoplasmic ratio is the oldest and best established model [1][2][9]. It proposes that suppressor molecules are present in the cytoplasm of the unfertilized egg and block several events like activation of zygotic gene expression [1]. During the first cell divisions, these hypothetical factors will be titrated out by the increasing number of nuclei relative to the constant total volume of cytoplasm. As soon as the concentration of repressing factors drops below a certain threshold, they will lose their repressing potential and MBT will start. This hypothesis has been supported by functional studies using nuclear transplantations and experimental manipulation of the cytoplasmic volume. Such experiments resulted either in a delayed or a premature beginning of the MBT [2][3]. Data from haploid [5] or tetraploid [2][5] animals strengthened these observations. Addition of extra DNA also led to an earlier start of the MBT [16][17][18]. Molecular, cellular and embryonic processes at early stages before MBT are neither well characterized nor fully understood for teleosts in general and medaka (Oryzias latipes) in particular. The medaka is a laboratory fish model of growing importance. It is comparable to zebrafish and also holds many features that legitimate it as a useful complementary model system [19]. For developmental studies, both model systems are of great interest since fertilization of eggs and embryonic development are external and embryos are totally transparent throughout the complete embryonic development. However, medaka may be more expedient than zebrafish for a couple of approaches that deal with the very early embryogenesis because of its slower embryogenesis. Zebrafish hatch after 2–3 days post fertilization, whereas medaka embryos develop much slower and do not hatch until day 7. While a lot of research aiming at MBT in fish in general has been performed using zebrafish [2] only one recent study PLoS ONE | www.plosone.org 1 July 2011 | Volume 6 | Issue 7 | e21741