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Zygotically transcribed genes
Large-scale movements of epithelial sheets are necessary for most embryonic and regenerative morphogenetic events. The cellular processes associated with germ band retraction (GBR) have been characterized in the Drosophila embryo. During GBR, the caudal end of the embryo retracts to its final posterior position. Using time-lapse recordings, it has been shown that, in contrast to germ band extension, cells within the lateral germ band do not intercalate. In addition, the germ band and amnioserosa move as one coherent sheet, and the amnioserosa strongly shortens along its dorsal-ventral axis. Furthermore, during GBR, the amnioserosa adheres to and migrates over the caudal end of the germ band via lamellipodia. Expression of both dominant-negative and constitutively active RhoA in the amnioserosa disrupts GBR. Since RhoA acts on both actomyosin contractility and cell-matrix adhesion, it suggests a role for such processes in the amnioserosa during GBR. The results establish the cellular movements and shape changes occurring during GBR and provide the basis for an analysis of the forces acting during GBR (Schock, 2002).
GBR is completed during embryonic stage 12. This is a time of exceptional morphogenetic activity: the midgut fuses and encloses the yolk sac laterally; the tracheal pit extensions fuse to form the tracheal tree, and the segmental furrows form from anterior to posterior. At stage 12, the embryo consists of two major epithelia, the squamous extraembryonic amnioserosa and the ectodermal germ band epithelium, as well as a mesenchymal mass of mesodermal and central nervous system precursor cells. Also found in the embryo are the epithelia of the foregut, hindgut, and salivary and tracheal pit invaginations, which are of ectodermal origin, and the midgut epithelium, which forms at that stage by mesenchymal to epithelial transition. The syncytial yolk sac, which is enclosed by a yolk sac membrane, sits in the middle of the embryo at the beginning of GBR, but moves more dorsally, directly beneath the amnioserosa, by the end of GBR. Analysis was focused on the amnioserosa and the germ band, because they appear to be the most likely candidates of the above tissues to participate in GBR. Cells in the germ band do not intercalate. GBR is therefore not a reversal of germ band extension (Schock, 2002).
Rather, reciprocal cell shape changes within the amnioserosa and the germ band are associated with the changes in embryo morphology at this stage. That is, the amnioserosa shortens along the DV axis, while the germ band elongates along the DV axis. This is possible because the amnioserosa and germ band are tightly attached to each other via adherens junctions, i.e., both epithelia move as one coherent sheet (Schock, 2002).
The boundary between amnioserosa and germ band was investigated at high magnification to obtain an idea of whether the shape changes observed are of an active or a passive nature. It was assumed that contractile forces would be exerted along the plasma membranes, because the actin cytoskeleton is localized cortically in both germ band and amnioserosa cells. The row of leading edge germ band cells is pulled in where the amnioserosa membranes perpendicular to the leading edge are attached. This suggests that amnioserosa cells contract along their DV axis. Cells of the germ band would be expected to push into the larger amnioserosa cells in the case of an active DV extension of the germ band, thus resulting in a convex shape of the amnioserosa-germ band boundary (Schock, 2002).
The presence of protrusions is demonstrated; these are formed predominantly at the posterior edge of the amnioserosa projecting toward the posterior. These protrusions exhibit high levels of dynamic actinGFP at the migration front, indicating actin polymerization. These protrusions have been classified as lamellipodia, because their appearance, behavior, and dynamic actin content are identical to lamellipodia in other motile cells. These lamellipodia migrate over the germ band instead of being passively dragged by the retracting germ band. The lamellipodia may migrate on an apical extracellular matrix secreted by germ band cells as a precursor to the larval cuticle. These observations indicate that the overlap of the amnioserosa over the caudal end of the germ band during GBR is maintained by lamellipodia-mediated migration. Furthermore, both constitutively active and dominant-negative RhoA disrupt GBR, when expressed in the amnioserosa. This suggests that actomyosin contractility or cell migration within the amnioserosa contribute to GBR, since these processes are affected by expression of rhoA mutants in tissue culture (Schock, 2002).
The amnioserosa is required for GBR because embryos that lack this tissue fail to undergo GBR. It has been proposed that this requirement for the amnioserosa may involve signaling from the amnioserosa to the germ band. The cell shape changes and motility observed within the amnioserosa and the overexpression experiments suggest that the amnioserosa additionally contributes to GBR in other ways than signaling (Schock, 2002).
The processes observed in this study allow several mechanisms, which are not mutually exclusive, to participate in GBR. (1) Segment furrow formation within the germ band may facilitate GBR by causing AP shortening of the germ band. (2) Active DV shortening of the amnioserosa may contribute to GBR by pulling in the lateral sides of the anterior germ band, thereby resulting in retraction of the germ band behind the bend of the U-shaped germ band-extended embryo. (3) The pulling force that appears to be exerted by DV shortening of the amnioserosa may be assisted by active DV extension of the germ band cells. (4) The overlap of the amnioserosa over the germ band may allow proper deployment of forces occurring within the amnioserosa (Schock, 2002).
Schock, F. and Perrimon, N. (2002). Cellular processes associated with germ band retraction in Drosophila. Dev. Bio. 248: 29-39. 12142018
date revised: 20 October 2002
Genes involved in organ development
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