Evolution of deuterostomy – and origin of the chordates

The chordates are usually characterized as bilaterians showing deuterostomy, i.e. the mouth developing as a new opening between the archenteron and the ectoderm, serial gill pores/slits, and the complex of chorda and neural tube. Both numerous molecular studies and studies of morphology and embryology demonstrate that the neural tube must be considered homologous to the ventral nerve cord(s) of the protostomes, but the origin of the ‘new’ mouth of the deuterostomes has remained enigmatic. However, deuterostomy is known to occur in several protostomian groups, such as the chaetognaths and representatives of annelids, molluscs, arthropods and priapulans. This raises the question whether the deuterostomian mouth is in fact homologous with that of the protostomes, viz. the anterior opening of the ancestral blastopore divided through lateral blastopore fusion, i.e. amphistomy. A few studies of gene expression show identical expression patterns around mouth and anus in protostomes and deuterostomes. Closer studies of the embryology of ascidians and vertebrates show that the mouth/stomodaeum differentiates from the anterior edge of the neural plate. Together this indicates that the chordate mouth has moved to the anterior edge of the blastopore, so that the anterior loop of the ancestral circumblastoporal nerve cord, which is narrow in the protostomes, has become indistinguishable. In the vertebrates, the mouth has moved further around the anterior pole to the ‘ventral’ side. The conclusion must be that the chordate mouth (and that of the deuterostomes in general) is homologous to the protostomian mouth and that the latest common ancestor of protostomes and deuterostomes developed through amphistomy, as suggested by the trochaea theory.     

Dorsoventral axis inversion: A phylogenetic perspective

Recent molecular evidence suggests that the body plans of insects and vertebrates may be dorsoventrally inverted with respect to one another. This poses a major challenge for comparative zoologists, either to explain how this came about or to offer alternative interpretations of the data. Dorsoventral inversion is most easily explained if the mouth of deuterostome metazoans (which would include vertebrates) is truly a secondary structure unrelated to the protostome mouth, located opposite to the latter on the dorsal surface of the body. Two possibilities are (1) that the definitive deuterostome mouth has replaced a preexisting protostome-type mouth, or (2) that ancestral deuterostomes lacked a protostome-type mouth altogether, and formed a secondary dorsal mouth entirely de novo. Our current understanding of invertebrate embryogenesis and larval morphology provides at least as much support, if not more, for (2) as for (1). By implication, even if ancestral protostomes and deuterostomes shared common anteroposterior and dorsoventral patterning systems, they may still have differed significantly with respect to other aspects of body organization and been otherwise very dissimilar organisms.     

Expression pattern of the Brachyury gene in the arrow worm paraspadella gotoi (chaetognatha)

Arrow worms (the phylum Chaetognatha), which are among the major marine planktonic animals, are direct developers and exhibit features characteristic of both deuterostomes and protostomes. In particular, the embryonic development of arrow worms appears to be of the deuterostome type. Brachyury functions critically in the formation of the notochord in chordates, whereas the gene is expressed in both the blastopore and stomodeum invagination regions in embryos of hemichordates and echinoderms. Here we analyzed the expression of Brachyury (Pg-Bra) in the arrow worm Paraspadella gotoi and showed that Pg-Bra is expressed in the blastopore region and the stomodeum region in the embryo and then around the mouth opening region at the time of hatching. The expression of Pg-Bra in the embryo resembles that of Brachyury in embryos of hemichordates and echinoderms, whereas that in the mouth opening region in the hatchling appears to be novel.     

Deuterostome evolution: early development in the enteropneust hemichordate, Ptychodera flava

SUMMARY Molecular and morphological comparisons indicate that the Echinodermata and Hemichordata represent closely related sister‐phyla within the Deuterostomia. Much less is known about the development of the hemichordates compared to other deuterostomes. For the first time, cell lineage analyses have been carried out for an indirect‐developing representative of the enteropneust hemichordates, Pty‐ chodera flava. Single blastomeres were iontophoretically labeled with DiI at the 2‐ through 16‐cell stages, and their fates followed through development to the tornaria larval stage. The early cleavage pattern of P. flava is similar to that of the direct‐developing hemichordate, Saccoglossus kowalevskii, as well as that displayed by indirect‐developing echinoids. The 16‐celled embryo contains eight animal “mesomeres,” four slightly larger “macromeres,” and four somewhat smaller vegetal “micromeres.” The first cleavage plane was not found to bear one specific relationship relative to the larval dorsoventral axis. Although individual blastomeres generate discrete clones of cells, the appearance and exact locations of these clones are variable with respect to the embryonic dorsoventral and bilateral axes. The eight animal mesomeres generate anterior (animal) ectoderm of the larva, which includes the apical organ; however, contributions to the apical organ were found to be variable as only a subset of the animal blastomeres end up contributing to its formation and this varies from embryo to embryo. The macromeres generate posterior larval ectoderm, and the vegetal micromeres form all the internal, endomesodermal tissues. These blastomere contributions are similar to those found during development of the only other hemichordate studied, the direct‐developing enteropneust, S. kowalevskii. Finally, isolated blastomeres prepared at either the two‐ or the four‐cell stage are capable of forming normal‐appearing, miniature tornaria larvae. These findings indicate that the fates of these cells and embryonic dorsoventral axial properties are not committed at these early stages of development. Comparisons with the developmental programs of other deuterostome phyla allow one to speculate on the conservation of some key developmental events/mechanisms and propose basal character states shared by the ancestor of echinoderms and hemichordates.     

The pericardium in the deuterostome <Emphasis Type="Italic">Saccoglossus kowalevskii</Emphasis> (Enteropneusta) develops from the ectoderm via schizocoely

The origin of mesoderm and coelomic compartments has traditionally been given high value for phylogenetic considerations of animal relationships. Two main modes have been distinguished, associated with the two main groups of animals: schizocoely with protostomes and enterocoely with deuterostomes. During enterocoely, coelomic compartments are formed from the endoderm. Here, we show that the pericardium of the deuterostome Saccoglossus kowalevskii, an enteropneust, is ontogenetically derived from the ectoderm and develops by schizocoely. The pericardium develops from a solid cluster of epidermis cells situated underneath the ectodermal nerve net above the basement membrane of the epidermis. The undifferentiated cells are interconnected by spot desmosomes, become separated from the epidermis and develop a central cavity. Pericardial cells become epithelial, by developing apical adherens junctions, a single apical cilium and basal striated myofibres. The differentiated pericardium possesses a cavity and surrounds a central blood vessel, the heart, situated in the basal extracellular matrix. The pericardium is an integral part of the anterior excretory complex, and comparisons to other deuterostomes indicate that pericardia are homologous despite differing ontogenies. Original data generated for the present study are deposited on MorphDBase ().     

Homeobox gene expression in Brachiopoda: the role of Not and Cdx in bodyplan patterning, neurogenesis, and germ layer specification

The molecular control that underlies brachiopod ontogeny is largely unknown. In order to contribute to this issue we analyzed the expression pattern of two homeobox containing genes, Not and Cdx, during development of the rhynchonelliform (i.e., articulate) brachiopod Terebratalia transversa. Not is a homeobox containing gene that regulates the formation of the notochord in chordates, while Cdx (caudal) is a ParaHox gene involved in the formation of posterior tissues of various animal phyla. The T. transversa homolog, TtrNot, is expressed in the ectoderm from the beginning of gastrulation until completion of larval development, which is marked by a three-lobed body with larval setae. Expression starts at gastrulation in two areas lateral to the blastopore and subsequently extends over the animal pole of the gastrula. With elongation of the gastrula, expression at the animal pole narrows to a small band, whereas the areas lateral to the blastopore shift slightly towards the future anterior region of the larva. Upon formation of the three larval body lobes, TtrNot expressing cells are present only in the posterior part of the apical lobe. Expression ceases entirely at the onset of larval setae formation. TtrNot expression is absent in unfertilized eggs, in embryos prior to gastrulation, and in settled individuals during and after metamorphosis. Comparison with the expression patterns of Not genes in other metazoan phyla suggests an ancestral role for this gene in gastrulation and germ layer (ectoderm) specification with co-opted functions in notochord formation in chordates and left/right determination in ambulacrarians and vertebrates. The caudal ortholog, TtrCdx, is first expressed in the ectoderm of the gastrulating embryo in the posterior region of the blastopore. Its expression stays stable in that domain until the blastopore is closed. Thereafter, the expression is confined to the ventral portion of the mantle lobe in the fully developed larva. No TtrCdx expression is detectable in the juvenile after metamorphosis. This expression of TtrCdx is congruent with findings in other metazoans, where genes belonging to the Cdx/caudal family are predominantly localized in posterior domains during gastrulation. Later in development this gene will play a fundamental role in the formation of posterior tissues.     

Origin of the chordate central nervous system – and the origin of chordates

 Contrary to traditional views, molecular evidence indicates that the protostomian ventral nerve cord plus apical brain is homologous with the vertebrates’ dorsal spinal cord plus brain. The origin of the protostomian central nervous system from a larval apical organ plus longitudinal areas along the fused blastopore lips has been documented in many species. The origin of the chordate central nervous system is more enigmatic. About a century ago, Garstang proposed that the ciliary band of a dipleurula-type larva resembling an echinoderm larva should have moved dorsally and fused to form the neural tube of the ancestral chordate. This idea is in contrast to a number of morphological observations, and it is here proposed that the neural tube evolved through lateral fusion of a ventral, postoral loop of the ciliary band in a dipleurula larva; the stomodaeum should move from the ventral side via the anterior end to the dorsal side, which faces the substratum in cephalo- chordates and vertebrates. This is in accordance with the embryological observations and with the molecular data on the dorsoventral orientation. The molecular observations further indicate that the anterior part of the insect brain is homologous with the anterior parts of the vertebrate brain. This leads to the hypothesis that the two organs evolved from the same area in the latest common bilaterian ancestor, just anterior to the blastopore, with the protostome brain developing from the anterior rim of the blastopore (i.e. in front of the protostome mouth) and the chordate brain from an area in front of the blastopore, but behind the mouth (i.e. behind the deuterostome mouth). Received: 28 August 1998 / Accepted: 14 November 1998     

Evolutionary modification of mouth position in deuterostomes

In chordates, the oral ectoderm is positioned at the anterior neural boundary and is characterized by pituitary homeobox (Pitx) and overlapping Dlx and Six3 expressions. Recent studies have shown that the ectoderm molecular map is also conserved in hemichordates and echinoderms. However, the mouth develops in a more posterior position in these animals, in a domain characterized by Nkx2.1 and Goosecoid expression, in a manner similar to that observed in protostomes. Furthermore, BMP signaling antagonizes mouth development in echinoderms and hemichordates, but seems to promote oral ectoderm specification in chordates. Conversely, Nodal signaling appears to be required for oral ectoderm specification in sea urchins but not in chordates. The Nodal/BMP antagonism at work during ectoderm patterning thus seems to constitute a conserved feature in deuterostomes, and mouth relocation may have been accompanied by a change in the influence of BMP/Nodal signals on oral ectoderm specification. We suggest that the mouth primordium was located at the anterior neural boundary, in early chordate evolution. In extant chordate embryos, subsequent mouth positioning differ between urochordates and vertebrates, presumably as a consequence of surrounding tissues remodelling. We illustrate these morphogenetic movements by means of morphological data obtained by the confocal imaging of ascidian tailbud embryos, and provide a table for determining the tailbud stages of this model organism.     

I-SceI Meganuclease-mediated transgenesis in the acorn worm, Saccoglossus kowalevskii

Hemichordates are a phylum of marine invertebrate deuterostomes that are closely related to chordates, and represent one of the most promising models to provide insights into early deuterostome evolution. The genome of the hemichordate, Saccoglossus kowalevskii, reveals an extensive set of non-coding elements conserved across all three deuterostome phyla. Functional characterization and cross-phyla comparisons of these putative regulatory elements will enable a better understanding of enhancer evolution, and subsequently how changes in gene regulation give rise to morphological innovation. Here, we describe an efficient method of transgenesis for the characterization of non-coding elements in S. kowalevskii. We first test the capacity of an I-SceI transgenesis system to drive ubiquitous or regionalized gene expression, and to label specific cell types. Finally, we identified a minimal promoter that can be used to test the capacity of putative enhancers in S. kowalevskii. This work demonstrates that this I-SceI transgenesis technique, when coupled with an understanding of chromatin accessibility, can be a powerful tool for studying how evolutionary changes in gene regulatory mechanisms contributed to the diversification of body plans in deuterostomes.     

Expression of a<Emphasis Type="Italic"> SoxB</Emphasis> and a<Emphasis Type="Italic"> Wnt2/13</Emphasis> gene during the development of the mollusc<Emphasis Type="Italic"> Patella vulgata</Emphasis>

We cloned and analysed the expression of a SoxB gene (PvuSoxB) in the marine mollusc, Patella vulgata. Like its orthologues in deuterostomes, after an early broad ectodermal distribution, PvuSoxB expression only persists in cells competent to form neural structures. In the post-gastrulation larva, PvuSoxB is expressed in the prospective neuroectoderm in the head and in the trunk. No expression can be seen dorsally, around the mouth and the anus, or along the ventral midline. We also report the expression of a Wnt2/13 orthologue (PvuWnt2) in Patella. After gastrulation, PvuWnt2 is expressed in the posterior part of the mouth, along the ventral midline and around the anus. This expression seems to be complementary to that of PvuSoxB in the trunk. We suggest the existence of a fundamental subdivision of the Patella trunk ectoderm into midline (mouth, midline, anus) and more lateral structures.Edited by D. Tautz     

Positioning the extreme anterior in Xenopus: cement gland, primary mouth and anterior pituitary

The extreme anterior of the deuterostome embryo is unusual in that ectoderm and endoderm are directly juxtaposed, without intervening mesoderm. In all vertebrates, this region gives rise to the anterior pituitary, the primary mouth and, in most frogs, to the mucus-secreting cement gland. Using the frog Xenopus laevis as a paradigm, we suggest that, initially, the extreme anterior forms a homogenous domain characterized by expression of pitx genes. Subsequently, this domain becomes subdivided to form these three different structures under the influence of different inductive signals from surrounding tissues.     

Observations et discussions sur le développement embryonnaire des Phoronida

Resume La fécondation est généralement interne chez les phoronidiens. La segmentation des ceufs est totale, egale (parfois légèrement inégale) et de type radiaire (avec quelquefois une apparence fortuite de segmentation spirale). La gastrula est formée par embolie. La bouehe derive de la zone blastoporale sans formation d'un vrai stomodeum. L'anus est mis en place par perforation de l'ectodersme et représente une néo-formation indépendante du blastopore. Le mesoderme est issu par proliferation cellulaire des regions antérieure et laterales de l'archentéron. Le protoccele est forme par des cellules mésodermiques se disposant le long de la paroi du lobe préoral. Le métaccele est issu probablement suivant les espèces d'une ou deux masses. La formation du mesoderme correspond á une variation de la méthode entéroccelique typique. Les phoronidiens doivent être considérés comme des deutérostomiens, d'après l'ensemble de nos résultats (voir aussi Emig, 1973).

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Observations and discussions on the embryonic Development in Phoronida

Summary Internal fertilization (in metaccelom) generally occurs in Phoronida. The eggs are extruded to the exterior through the nephridia, shed freely into the sea-water or retained in the lophophoral concavity. The cleavage of phoronid eggs is total, equal (or subequal) and radial (with sometimes fortuitous appearance of spiral cleavage patterns). The gastrula is formed by emboly. The mouth is derived from the anterior remnant of the blastopore without a true stomodeum. The anus arises by perforation, as an independent structure of the blastopore. The mesoderm formed by budding originates as isolated cells proliferated from the anterior and lateral surfaces of the archenteron. In the preoral hood appears a protoccel by mesodermal cells lining the walls of the blastoccel. The trunk clom (or metaccel) of Actinotrocha originates from one or two posterior masses of mesodermal cells. It is possible that the mode of formation of this coelom varies in respect to the different species. The mesoderm elaboration is considered as a modified enteroccelous method.The acceptance of Phoronida as deuterostomes is regarded as the logical consequence of the present considerations (see also Emig, 1973): radial cleavage, origin of mesoderm by a derived enteroccelous method, trimetamerous actinotrocha.

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Abbréviations des figures a anus - lp lobe préoral - ar archentéron - b blastoccele - ma mésoderme de la région anterieure de l'archentéron - ml mésoderme des régions latérales de l'archentéron - bl blastopore - mes cellule mésodermique - bo bouche - n ebauche des - n éphridiesect mesderme - s sophage - end endoderme - p protocle - est estomac - t ebauche des tentacules - g ebauche du ganglion nerveux - te tentacule - gn glandes nidamentaire - v vestibule - i intestin     

Furrowing surface contraction wave coincident with primary neural induction in amphibian embryos

We predicted, and have now observed, a surface contraction wave in axolotl (Ambystoma mexicanum) embryos that appears to coincide temporally and spatially with primary neural induction and homoiogenetic induction, and with involution of the chordomesoderm. The wave starts from a focus anterior to the dorsal lip of the blastopore and spreads as an ellipse, until part of it encounters the rim of the blastopore and vanishes there. The remaining are then continues over the dorsal hemisphere until it reforms an ellipse that decreases in size. About 9 to 12 hours after it begins, the wave vanishes at a focus diametrically opposite its point of origin. The wave involves both local contraction and furrowing in the monolayer ectoderm. To a good approximation, the hemispherical portion of the ectoderm traversed by the wave becomes neuroepithelium, while the ectoderm not transversed by the wave becomes epidermis. The wave might provide a mechanism to determine the time and location at which neuroepithelial differentiation occurs. © 1994 Wiley-Liss, Inc.     

Ventralization of an indirect developing hemichordate by NiCl2 suggests a conserved mechanism of dorso-ventral (D/V) patterning in Ambulacraria (hemichordates and echinoderms)

One of the earliest steps in embryonic development is the establishment of the future body axes. Morphological and molecular data place the Ambulacraria (echinoderms and hemichordates) within the Deuterostomia and as the sister taxon to chordates. Extensive work over the last decades in echinoid (sea urchins) echinoderms has led to the characterization of gene regulatory networks underlying germ layer specification and axis formation during embryogenesis. However, with the exception of recent studies from a direct developing hemichordate (Saccoglossus kowalevskii), very little is known about the molecular mechanism underlying early hemichordate development. Unlike echinoids, indirect developing hemichordates retain the larval body axes and major larval tissues after metamorphosis into the adult worm. In order to gain insight into dorso-ventral (D/V) patterning, we used nickel chloride (NiCl₂), a potent ventralizing agent on echinoderm embryos, on the indirect developing enteropneust hemichordate, Ptychodera flava. Our present study shows that NiCl₂ disrupts the D/V axis and induces formation of a circumferential mouth when treated before the onset of gastrulation. Molecular analysis, using newly isolated tissue-specific markers, shows that the ventral ectoderm is expanded at expense of dorsal ectoderm in treated embryos, but has little effect on germ layer or anterior–posterior markers. The resulting ventralized phenotype, the effective dose, and the NiCl₂ sensitive response period of Ptychodera flava, is very similar to the effects of nickel on embryonic development described in larval echinoderms. These strong similarities allow one to speculate that a NiCl₂ sensitive pathway involved in dorso-ventral patterning may be shared between echinoderms, hemichordates and a putative ambulacrarian ancestor. Furthermore, nickel treatments ventralize the direct developing hemichordate, S. kowalevskii indicating that a common pathway patterns both larval and adult body plans of the ambulacrarian ancestor and provides insight in to the origin of the chordate body plan.     

Development of the primary mouth in Xenopus laevis

The initial opening between the gut and the outside of the deuterostome embryo breaks through at the extreme anterior. This region is unique in that ectoderm and endoderm are directly juxtaposed, without intervening mesoderm. This opening has been called the stomodeum, buccopharyngeal membrane or oral cavity at various stages of its formation, however, in order to clarify its function, we have termed this the "primary mouth". In vertebrates, the neural crest grows around the primary mouth to form the face and a "secondary mouth" forms. The primary mouth then becomes the pharyngeal opening. In order to establish a molecular understanding of primary mouth formation, we have begun to examine this process during Xenopus laevis development. An early step during this process occurs at tailbud and involves dissolution of the basement membrane between the ectoderm and endoderm. This is followed by ectodermal invagination to create the stomodeum. A subsequent step involves localized cell death in the ectoderm, which may lead to ectodermal thinning. Subsequently, ectoderm and endoderm apparently intercalate to generate one to two cell layers. The final step is perforation, where (after hatching) the primary mouth opens. Fate mapping has defined the ectodermal and endodermal regions that will form the primary mouth. Extirpations and transplants of these and adjacent regions indicate that, at tailbud, the oral ectoderm is not specifically required for primary mouth formation. In contrast, underlying endoderm and surrounding regions are crucial, presumably sources of necessary signals. This study indicates the complexity of primary mouth formation, and lays the groundwork for future molecular analyses of this important structure.