RESULTS

Cells are arranged into species-specific patterns during early embryogenesis. Such cell division patterns are important since they often reflect the distribution of localized cortical factors from eggs/fertilized eggs to specific cells as well as the emergence of organismal form. However, it has proven difficult to reveal the mechanisms that underlie the emergence of cell positioning patterns that underlie embryonic shape, likely because a systems-level approach is required that integrates cell biological, genetic, developmental, and mechanical parameters. The choice of organism to address such questions is also important. Because ascidians display the most extreme form of invariant cleavage pattern among the metazoans, we have been analyzing the cell biological mechanisms that underpin three aspects of cell division (unequal cell division (UCD), oriented cell division (OCD), and asynchronous cell cycles) which affect the overall shape of the blastula-stage ascidian embryo composed of 64 cells. In ascidians, UCD creates two small cells at the 16-cell stage that in turn undergo two further successive rounds of UCD. Starting at the 16-cell stage, the cell cycle becomes asynchronous, whereby the vegetal half divides before the animal half, thus creating 24-, 32-, 44-, and then 64-cell stages. Perturbing either UCD or the alternate cell division rhythm perturbs cell position. We propose that dynamic cell shape changes propagate throughout the embryo via cell-cell contacts to create the ascidian-specific invariant cleavage pattern

Embryos of the viviparous dermapteran, Arixenia esau develop sequentially in two compartments: terminal ovarian follicles and the uterus.

Three main reproductive strategies have been described among insects: most common oviparity, ovoviviparity and viviparity. In the latter strategy, the embryonic development takes place within the body of the mother which provides gas exchange and nutrients for embryos. Here we present the results of histological and EM analyses of the female reproductive system of the viviparous earwig, Arixenia esau, focusing on all the modifications related to the viviparity. We show that in the studied species the embryonic development consists of two "physiological phases" that take place in two clearly disparate compartments, i.e. the terminal ovarian follicle and the uterus. In both compartments the embryos are associated with synthetically active epithelial cells. We suggest that these cells are involved in the nourishment of the embryo. Our results indicate that viviparity in arixeniids is more complex than previously considered. We propose the new term "pseudoplacento-uterotrophic viviparity" for this unique two-phase reproductive strategy

Balbiani body of basal insects is potentially involved in multiplication and selective elimination of mitochondria

Abstract Oocytes of both vertebrates and invertebrates often contain an intricate organelle assemblage, termed the Balbiani body (Bb). It has previously been suggested that this assemblage is involved in the delivery of organelles and macromolecules to the germ plasm, formation of oocyte reserve materials, and transfer of mitochondria to the next generation. To gain further insight into the function of the Bb, we performed a series of analyses and experiments, including computer-aided 3-dimensional reconstructions, detection of DNA (mtDNA) synthesis as well as immunolocalization studies. We showed that in orthopteran Meconema meridionale, the Bb comprises a network of mitochondria and perinuclear nuage aggregations. As oogenesis progresses, the network expands filling almost entire ooplasm, then partitions into several smaller entities, termed micro-networks, and ultimately into individual mitochondria. As in somatic cells, this process involves microfilaments and elements of endoplasmic reticulum. We showed also that at least some of the individual mitochondria are surrounded by phagophores and eliminated via mitophagy. These findings support the idea that the Bb is implicated in the multiplication and selective elimination of (defective) mitochondria and therefore may participate in the transfer of undamaged (healthy) mitochondria to the next generation

Secretory activity of the uterus epithelial cells.

<p>Transverse section through the infolding of the uterus wall (boxed fragment of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0064087#pone-0064087-g001" target="_blank">Figure 1C</a>). Two closely opposed epithelial layers are visible. Note argyrophylic secretory vacuoles in the cytoplasm of epithelial cells (encircled). Basement lamina (empty arrows), tracheal branches (black arrowheads), argyrophylic material on the cell surface (white asterisks). Semi-thin section stained with AgNOR technique. LM, scale bar: 24 µm.</p

Embryonic development.

<p>(<b>A</b>) Fragment of a young embryo (cross section; blastoderm stage) developing inside a terminal ovarian follicle. DAPI staining. Note that the nuclei of follicular cells (fc) are substantially larger and “brighter” than blastoderm nuclei (bl). Yolk nuclei (yn). FM, scale bar: 24 µm. (<b>B, C</b>) Fragment of an older embryo (cross section; germ band stage) inside a terminal follicle. Note the germ band (gb) surrounded by the follicular epithelium (fc) and two embryonic envelopes, serosa (s) and amnion (a). The follicular epithelium is separated from the embryo by a thin structureless eggshell (arrowheads). LM, scale bar: 24 µm. (<b>D</b>) Posterior part of the embryo dissected from the lumen of the uterus. Note well developed cerci (ce) and the last abdominal segment (telson, t). SEM, scale bar: 0.5 mm. (<b>E</b>) Follicular cell (fragment) surrounding the blastoderm stage embryo. Note highly folded envelope surrounding irregularly shaped nucleus (n) and large secretory vacuoles (v). TEM, scale bar: 1 µm.</p

Morphology of the female reproductive system.

<p>(<b>A</b>) Three ovarioles (ov) attached to the uterus (u). White arrow indicates circumferential muscle fibers surrounding the posteriormost section of the uterus. Tracheal branches (black arrowheads). SEM, scale bar: 0.5 mm. (<b>B, C</b>) Transverse section through the uterus (fragment). Note folded epithelium (ep) lining the uterus (u). Basement lamina (empty arrows), muscle fibers (white arrowheads), tracheal branches (black arrowheads), grooves separating ridges of the epithelium (arrows), black asterisks in (C) indicate argyrophylic material on the cell surface. Boxed fragment in (C) is enlarged in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0064087#pone-0064087-g002" target="_blank">Figure 2</a>. (B) Semi-thin section stained with methylene blue. (C) Semi-thin section stained with AgNOR technique. LM, scale bar: 24 µm. (<b>D</b>) Basement lamina (bl) supporting epithelial cells (ep) of the uterus. The lamina is penetrated by canals (c) containing tracheal branches (black arrowheads) immersed in filamentous material. TEM, scale bar: 1 µm. (<b>E, F</b>) Apical compartment of an epithelial cell lining the uterus. Note large secretory vacuoles (v), fibro-granular material covering the tips of the microvilli (white asterisk) and an adherens junction (aj) connecting membranes of neighboring cells. Mitochondria (m). TEM, scale bar: 1 µm. (<b>G</b>) Basal compartments of epithelial cells of the uterus. Mitochondria (m), elements of rough endoplasmic reticulum (rer). Note that plasma membranes are folded and closely adjoined. TEM, scale bar: 1 µm.</p

Embryo after dorsal flexion of the germ band.

<p>(A) Longitudinal section through a central region of the embryo; note the egg cytoplasm (egc) surrounded by the germ band (gb); yolk cell nucleus (yn), margins of the germ band are indicated by double arrows, the wall of the embryonic follicle (ew); semithin section stained with methylene blue. Scale bar: 100 μm. (B) Individual yolk cell comprising ER stacks (arrows); yolk cell nucleus (yn); Nomarski interference contrast. Scale bar: 50 μm. (C) TEM micrograph showing dispersing ER stacks; note that marginally located ER elements (arrowheads) are partly separated from the stack (ERst) and covered with ribosomes. Scale bar: 1 μm. (D) Canonical ER network in association with ER stack (ERst); vacuoles (v); TEM micrograph. Scale bar: 1 μm.</p