The control of the anteroposterior and dorsoventral axes in embryonic chick limbs constructed of dissociated and reaggregated limb-bud mesoderm

In the 3- to 4-day embryonic avian limb bud, a unique zone of mesodermal tissue is located posteriorly at the junction of bud and body wall. Appropriately grafted to a host limb bud, it induces the formation of a supernumerary limb outgrowth from preaxial tissue and determines that its posterior side will face the graft. It is called the zone of polarizing activity (ZPA).When limb-bud mesoderm is isolated, dissociated, reaggregated centrifugally, jacketed in the mesoderm-free hull of another limb bud, and grown as a graft on a host embryo, the recombinant frequently forms a limb-like structure terminating in digits that fail to show differentiation with respect to the anteroposterior axis. When, however, a bit of ZPA tissue is implanted in the recombinant subjacent to the anterior or posterior margin of the ectoderm, the resulting outgrowth shows a characteristic anteroposterior order of digits that corresponds to the placement of the implant, regardless of its relationship with the anteroposterior axis of the ectoderm or of the host embryo.Dorsoventral differentials have been recognized only in limbs formed from reaggregated leg-bud mesoderm. The direction of the dorsoventral axis always corresponds to the original axis of the ectodermal jacket regardless of the orientation of the recombinant on the host.     

The anterior-posterior axis emerges respecting the morphology of the mouse embryo that changes and aligns with the uterus before gastrulation

BACKGROUND: When the anterior-posterior axis of the mouse embryo becomes explicit at gastrulation, it is almost perpendicular to the long uterine axis. This led to the belief that the uterus could play a key role in positioning this future body axis. RESULTS: Here, we demonstrate that when the anterior-posterior axis first emerges it does not respect the axes of the uterus but, rather, the morphology of the embryo. Unexpectedly, the emerging anterior-posterior axis is initially aligned not with the long, but the short axis of the embryo. Then whether the embryo develops in vitro or in utero, the anterior-posterior axis becomes aligned with the long axis of embryo just prior to gastrulation. Of three mechanisms that could account for this apparent shift in anterior-posterior axis orientation-cell migration, spatial change of gene expression, or change in embryo shape-lineage tracing studies favor a shape change accompanied by restriction of the expression domain of anterior markers. This property of the embryo must be modulated by interactions with the uterus as ultimately the anterior-posterior and long axes of the embryo align with the left-right uterine axis. CONCLUSIONS: The emerging anterior-posterior axis relates to embryo morphology rather than that of the uterus. The apparent shift in its orientation to align with the long embryonic axis and with the uterus is associated with a change in embryo shape and a refinement of anterior gene expression pattern. This suggests an interdependence between anterior-posterior gene expression, the shape of the embryo, and the uterus.     

The polarising role of cell adhesion molecules in early development

Polarising a cell or an embryo is a crucial and recurrent event during development, as it is important for cell differentiation and migration. Cells can become polarised along their apical-basal axis and also within the plane of the tissue layer to which they belong. The embryo develops three axes: the anteroposterior, the dorsoventral and the left-right axis. Recent work indicates instructive roles for cell adhesion molecules in establishing not only apical-basal polarity but also planar cell polarity and, surprisingly, in the generation of left-right asymmetry in vertebrates. Signalling cascades that regulate polarity formation seem to be conserved among different organisms, thereby raising the intriguing question of whether this also holds true for the cell adhesion molecules.     

Secondary embryonic axis formation by transplantation of D quadrant micromeres in an oligochaete annelid

Among spiral cleaving embryos (e.g. mollusks and annelids), it has long been known that one blastomere at the four-cell stage, the D cell, and its direct descendants play an important role in axial pattern formation. Various studies have suggested that the D quadrant acts as the organizer of the embryonic axes in annelids, although this has never been demonstrated directly. Here we show that D quadrant micromeres (2d and 4d) of the oligochaete annelid Tubifex tubifex are essential for embryonic axis formation. When 2d and 4d were ablated the embryo developed into a rounded cell mass covered with an epithelial cell sheet. To examine whether 2d and 4d are sufficient for axis formation they were transplanted to an ectopic position in an otherwise intact embryo. The reconstituted embryo formed a secondary embryonic axis with a duplicated head and/or tail. Cell lineage analyses showed that neuroectoderm and mesoderm along the secondary axis were derived from the transplanted D quadrant micromeres and not from the host embryo. However, endodermal tissue along the secondary axis originated from the host embryo. Interestingly, when either 2d or 4d was transplanted separately to host embryos, the reconstituted embryos failed to form a secondary axis, suggesting that both 2d and 4d are required for secondary axis formation. Thus, the Tubifex D quadrant micromeres have the ability to organize axis formation, but they lack the ability to induce neuroectodermal tissues, a characteristic common to chordate primary embryonic organizers.     

Strcture of the Mature Embryo of Theobroma cacao L.

The mature embryo of Theobroma cacao L. consists of an embryonicaxis and two cotyledons. The cotyledonary tissue comprises thebulk of the embryo and contains mature and maturing vasculartissue surrounded by a bundle sheath. The cells of the groundtissue are filled with starch grains, anthocyanin pigments,or mucilage. The embryonic axis consists of epicotyl, hypocotyl,and root meristem. Maturation of the vascular tissue has begunin the procambium, where xylem maturation is ahead of phloemmaturation. Mucilage cells are found throughout the tissue of the axis.Most of the cells of the embryonic tissue contain starch grains.     

Induction and migration of the anterior visceral endoderm is regulated by the extra-embryonic ectoderm

The anterior visceral endoderm (AVE) is an extra-embryonic tissue required for specifying anterior pattern in the mouse embryo. The AVE is induced at the distal tip of the 5.5 dpc embryo and then migrates to the prospective anterior, where it imparts anterior identity upon the underlying epiblast (the tissue that gives rise to the embryo proper). Little is known about how the AVE is induced and what directs its migration. In this paper, we describe an essential role for another extra-embryonic tissue, the extra-embryonic ectoderm (ExE), in patterning the AVE and epiblast. Removal of the ExE in pre-gastrulation embryos leads to ectopic AVE formation, to a failure of AVE cell migration and to the assumption by the entire epiblast of an anterior identity. Ectopic transplantation of ExE cells inhibits AVE formation and leads to an expansion of the posterior epiblast marker T. These results demonstrate that the ExE restricts the induction of the AVE to the distal tip of the mouse embryo and is required to initiate the migration of these cells to the prospective anterior. Together, these data reveal a novel role for the ExE in the specification of the anteroposterior axis of the mouse embryo.     

Polarity of the mouse embryo is anticipated before implantation

In most species, the polarity of an embryo underlies the future body plan and is determined from that of the zygote. However, mammals are thought to be an exception to this; in the mouse, polarity is generally thought to develop significantly later, only after implantation. It has not been possible, however, to relate the polarity of the preimplantation mouse embryo to that of the later conceptus due to the lack of markers that endure long enough to follow lineages through implantation. To test whether early developmental events could provide cues that predict the axes of the postimplantation embryo, we have used the strategy of injecting mRNA encoding an enduring marker to trace the progeny of inner cell mass cells into the postimplantation visceral endoderm. This tissue, although it has an extraembryonic fate, plays a role in axis determination in adjacent embryonic tissue. We found that visceral endoderm cells that originated near the polar body (a marker of the blastocyst axis of symmetry) generally became distal as the egg cylinder formed, while those that originated opposite the polar body tended to become proximal. It follows that, in normal development, bilateral symmetry of the mouse blastocyst anticipates the polarity of the later conceptus. Moreover, our results show that transformation of the blastocyst axis of symmetry into the axes of the postimplantation conceptus involves asymmetric visceral endoderm cell movement. Therefore, even if the definitive axes of the mouse embryo become irreversibly established only after implantation, this polarity can be traced back to events before implantation.     

蓖麻生物工程研究进展

蓖麻是一种高蓄能植物和工业原料植物,具有很大的开发利用价值。本文从组织培养和遗传转化两个方面并结合本实验室的工作综述了蓖麻生物工程研究的最新进展。在蓖麻组织培养方面,不同的外植体中以成熟种子的胚轴最适宜,而在不同激素中以TDZ诱导丛生芽的效率最高,并以此为基础建立了蓖麻离体再生体系。在遗传转化方面,不同的转化方法中以农杆菌介导法最适合蓖麻转化。蓖麻胚轴对卡那霉素不敏感,潮霉素是蓖麻转化的适宜筛选剂。文中指出了蓖麻生物工程研究中存在的问题,并对应用生物技术培育蓖麻新品种和促进蓖麻生产的可能性进行了讨论。     

Active cell migration drives the unilateral movements of the anterior visceral endoderm

The anterior visceral endoderm (AVE) of the mouse embryo is a specialised extra-embryonic tissue that is essential for anterior patterning of the embryo. It is characterised by the expression of anterior markers such as Hex, Cerberus-like and Lhx1. At pre-gastrula stages, cells of the AVE are initially located at the distal tip of the embryo, but they then move unilaterally to the future anterior. This movement is essential for converting the existing proximodistal axis into an anteroposterior axis. To investigate this process, we developed a culture system capable of imaging embryos in real time with single cell resolution. Our results show that AVE cells continuously change shape and project filopodial processes in their direction of motion, suggesting that they are actively migrating. Their proximal movement stops abruptly at the junction of the epiblast and extra-embryonic ectoderm, whereupon they move laterally. Confocal microscope images show that AVE cells migrate as a single layer in direct contact with the epiblast, suggesting that this tissue might provide directional cues. Together, these results show that the anteroposterior axis is correctly positioned by the active movement of cells of the AVE in response to cues from their environment, and by a 'barrier' to their movement that provides an endpoint for this migration.     

Spatial distribution of the capacity to initiate a secondary embryo in the 32-cell embryo of Xenopus laevis

To examine the spatial distribution of dorsal determinants in the early embryos of Xenopus laevis, individual cells from the 32-cell embryo were transplanted into the same tier of the ventral side of a synchronous recipient. Their abilities to initiate a secondary embryo were measured by the incidence of secondary embryos and by the length of the secondary axis relative to the primary embryo. The ability was found to be localized in all cells (A1, B1, C1, and D1) of the dorsal most column and in the vegetal cells (C2 and D2) of the dorsolateral column. Transplanted C1 (subequatorial) cells caused the highest incidence of a secondary embryo and the average relative length of the secondary embryo was also greatest. Effectiveness decreased in the order: D1, B1, D2, C2, and A1. When these results were compared with Dale and Slack's fate map of the 32-cell embryo, it was concluded that the distribution of dorsal determinants is unique and does not coincide with the prospective regions for any tissues, though it is somewhat similar to the prospective region of dorsal endoderm or notochord. From these results it seems that dorsal determinants do not determine a particular tissue in an embryo but rather the "dorsal" region of an embryo.     

Pattern formation in the flowering plant embryo.

Recent mutation studies in Arabidopsis suggest rules by which the primary organization of the plant body is established in the early embryo. The main types of plant tissue arise independently of pattern formation along the axis of polarity. The axis is initially partitioned into three regions. This prepattern is later refined, possibly by position-specific cell activities, as indicated by morphological features as well as the distribution of molecular markers.     

Morphogenetic tissue movement and the establishment of body plan during development from blastocyst to gastrula in the mouse

In many animal species, the early development of the embryo follows a stereotypic pattern of cell cleavage, lineage allocation and generation of tissue asymmetry leading to delineation of the body plan with three primary embryonic axes. The mammalian embryo has been regarded as an exception and primary body axes of the mouse embryo were thought to develop after implantation. However, recent findings have challenged this view. Asymmetry in the fertilised oocyte, as defined by the position of the second polar body and the sperm entry point, can be correlated with the orientation of the animal-vegetal and the embryonic-abembryonic axes in the preimplantation blastocyst. Studies of the pattern of morphogenetic movement of cells and genetic activity in the peri-implantation embryo suggest that the animal-vegetal axis of the blastocyst might presage the orientation of the anterior-posterior axis of the gastrula. This suggests that the asymmetry of the zygote that is established at fertilisation and early cleavage has a lasting impact on the delineation of body axes during embryogenesis.     

The anterior visceral endoderm-turning heads

The Anterior Visceral Endoderm is an extraembryonic tissue that plays a pivotal role during embryogenesis, being responsible for the proper orientation of the anterior-posterior axis of the embryo and for appropriate pattering of adjacent embryonic tissue. In this review I discuss the formation and migration of the AVE, and attempt to place some recent findings in the context of a working model.     

Control of early anterior-posterior patterning in the mouse embryo by TGF-beta signalling

Prior to gastrulation the mouse embryo exists as a symmetrical cylinder consisting of three tissue layers. Positioning of the future anterior-posterior axis of the embryo occurs through coordinated cell movements that rotate a pre-existing proximal-distal (P-D) axis. Overt axis formation becomes evident when a discrete population of proximal epiblast cells become induced to form mesoderm, initiating primitive streak formation and marking the posterior side of the embryo. Over the next 12-24 h the primitive streak gradually elongates along the posterior side of the epiblast to reach the distal tip. The most anterior streak cells comprise the 'organizer' region and include the precursors of the so-called 'axial mesendoderm', namely the anterior definitive endoderm and prechordal plate mesoderm, as well as those cells that give rise to the morphologically patent node. Signalling pathways controlled by the transforming growth factor-beta ligand nodal are involved in orchestrating the process of axis formation. Embryos lacking nodal activity arrest development before gastrulation, reflecting an essential role for nodal in establishing P-D polarity by generating and maintaining the molecular pattern within the epiblast, extraembryonic ectoderm and the visceral endoderm. Using a genetic strategy to manipulate temporal and spatial domains of nodal expression reveals that the nodal pathway is also instrumental in controlling both the morphogenetic movements required for orientation of the final axis and for specification of the axial mesendoderm progenitors.     

Control by the Embryo Axis of the Breakdown of Storage Proteins in the Endosperm of Germinated Castor BeanSeed: A Role for Gibberellic Acid

The embryo axis is required for the rapid breakdown of the crystalloid,albumin and lectin protein storage reserve in the endospermof castor bean (Ricinus communis L. cv. Hale) seeds, and forthe attainment of high specific activities of several endospermicproteolytic enzymes: one carboxy-peptidase and two -SH- dependentaminopeptidases. The embryo axis must be present to initiatestorage protein breakdown but it is not required to maintainthis process. We suggest that the embryo axis controls storageprotein breakdown through the release of promoters, which canbe replaced by gibberellins. Storage protein breakdown is notinfluenced by source-sink effects. However, the endosperm becomessensitive to gibberellin only after an imbibition period forup to 24 h. Key words: Castor bean, Protein breakdown, Storage protein, Embryo control, Gibberellin, Seed germination     

Differences in the activity and distribution of peroxidases from three different portions of germinating Brassica oleracea seeds

Peroxidase (POD, EC 1.11.1.7) activity, cellular localization and isozyme patterns were investigated in the seed integument, cotyledon and embryo axis of Brassica oleracea cv. Cappuccio during pregermination and seedling growth. Seeds started to germinate after 24 h of imbibition. POD activity was localized in the pigmented layer of the integument and in procambial strands of the cotyledon and embryo axis in the first 24 h of imbibition. It was localized in the integumental cells of palisade, pigmented and aleurone layers and in epidermal, meristematic, procambial cells and xylem elements of the root and hypocotyl after 48 h of imbibition. POD activity increased during germination and early seedling growth: in the integument, it reached a maximum value after 72 h of imbibition, in the embryo axis and cotyledons, it increased up to 144 h of imbibition. The increase in peroxidase activity was accompanied by the appearance of new isozymes correlated with the development of seedling tissues. The isozyme profile was characterized by nine peroxidases: isoperoxidase of 50 kDa peculiar to integuments, that of 150 kDa to cotyledons and that of 82 kDa to the embryo axis. During pregerminative phase isozymes of 84 kDa were detected in the integument and cotyledons, of 48.5 kDa in the embryo axis. After germination, peroxidase activity and the complexity of the isozyme pattern increased, suggesting that they play a relevant role after rupture of the integument.     

Effect of plumule and radicle on somatic embryogenesis in the cultures of ginseng zygotic embryos

Immature zygotic embryos of ginseng produced somatic embryos on MS medium without growth regulators. However, in the culture of mature zygotic embryos, excision of the embryo was required for somatic embryo induction. Somatic embryos formed only on excised cotyledons without an embryo axis or on excised embryos without the plumule and radicle of the axis. This observation suggests that the axis tip of the embryo might suppress somatic embryo production although the cotyledon tissues have predetermined embryogenic competency. To clarify the role of the embryo axis on somatic embryo formation, excised plumules or radicles were placed in direct contact with the basal cut-ends of cotyledons. The adhesion of plumules or radicles highly suppressed somatic embryo formation from cotyledon explants. When an agar block containing exudate from excised plumules or radicles was placed in contact with the cut end of the cotyledon, a similar inhibition was observed. These results suggest that embryogenic competence is suppressed by endogenous inhibitors present in the axis tip of the zygotic embryo.     

The first cleavage furrow demarcates the dorsal-ventral axis in Xenopus embryos

To determine the relationship between the first cleavage furrow and the dorsal-ventral axis of the Xenopus embryo, a heritable intracellular marker was injected into one blastomere at the two-cell stage. Embryos were selected in which the cleavage furrow bisected the crescent-shaped region of pale pigmentation or in which it formed 45-90 degrees from this region. This region, which is located in the animal hemisphere of the Xenopus embryo, meets the criteria of the grey crescent as defined in other amphibian species. At tailbud stages the interface between the labeled and unlabeled halves was always coincident with the midsagittal plane. This correlation shows that the first cleavage furrow demarcates the dorsal-ventral axis. The labeling pattern was the same whether the first cleavage furrow bisected the region of pale pigmentation or whether it formed 90 degrees from it. However, when this region was bisected (70% of embryos) it always was located on the dorsal side of the embryo. Thus the region of pale pigmentation indicates the dorsal side of the embryo only when it is bisected by the first cleavage furrow.