Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 4 (9), e291

Dorsoventral Patterning in Hemichordates: Insights Into Early Chordate Evolution

Affiliations

Dorsoventral Patterning in Hemichordates: Insights Into Early Chordate Evolution

Christopher J Lowe et al. PLoS Biol.

Erratum in

Abstract

We have compared the dorsoventral development of hemichordates and chordates to deduce the organization of their common ancestor, and hence to identify the evolutionary modifications of the chordate body axis after the lineages split. In the hemichordate embryo, genes encoding bone morphogenetic proteins (Bmp) 2/4 and 5/8, as well as several genes for modulators of Bmp activity, are expressed in a thin stripe of ectoderm on one midline, historically called "dorsal." On the opposite midline, the genes encoding Chordin and Anti-dorsalizing morphogenetic protein (Admp) are expressed. Thus, we find a Bmp-Chordin developmental axis preceding and underlying the anatomical dorsoventral axis of hemichordates, adding to the evidence from Drosophila and chordates that this axis may be at least as ancient as the first bilateral animals. Numerous genes encoding transcription factors and signaling ligands are expressed in the three germ layers of hemichordate embryos in distinct dorsoventral domains, such as pox neuro, pituitary homeobox, distalless, and tbx2/3 on the Bmp side and netrin, mnx, mox, and single-minded on the Chordin-Admp side. When we expose the embryo to excess Bmp protein, or when we deplete endogenous Bmp by small interfering RNA injections, these expression domains expand or contract, reflecting their activation or repression by Bmp, and the embryos develop as dorsalized or ventralized limit forms. Dorsoventral patterning is independent of anterior/posterior patterning, as in Drosophila but not chordates. Unlike both chordates and Drosophila, neural gene expression in hemichordates is not repressed by high Bmp levels, consistent with their development of a diffuse rather than centralized nervous system. We suggest that the common ancestor of hemichordates and chordates did not use its Bmp-Chordin axis to segregate epidermal and neural ectoderm but to pattern many other dorsoventral aspects of the germ layers, including neural cell fates within a diffuse nervous system. Accordingly, centralization was added in the chordate line by neural-epidermal segregation, mediated by the pre-existing Bmp-Chordin axis. Finally, since hemichordates develop the mouth on the non-Bmp side, like arthropods but opposite to chordates, the mouth and Bmp-Chordin axis may have rearranged in the chordate line, one relative to the other.

Conflict of interest statement

Competing interests. The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. The Anatomy of S. kowalevskii to Illustrate Differentiations along the Dorsoventral Axis
The ventral side, assigned because of the mouth location, is down in this schematic figure. Anterior is to the left. Note the three parts of the body: the prosome (proboscis) at the anterior end, the narrow mesosome (collar) in the middle, and the elongated metasome to the posterior (only half the length is shown). Ectodermal derivatives are colored blue, mesodermal red, and endodermal yellow. Prominent structures of the dorsoventral axis include the dorsal and ventral axon tracts, the dorsal and ventral blood vessels, the dorsal stomochord with the heart/kidney complex, the dorsal gill slits, the ventral mouth, and the ventral tail (only in the juvenile, not shown). Redrawn with modification from Benito and Pardos [105].
Figure 2
Figure 2. Early Midline Signaling of Bmp and Chordin
All embryos are cleared (see Materials and Methods) otherwise specified. All embryos have a similar orientation: anterior in the top left and dorsal in the top right of each panel, unless otherwise specified. (A) Expression of bmp2/4 at late gastrula stage and (B) at day 3 of development. (C) Hatched juvenile oriented with dorsal toward the viewer. White arrowhead indicates the position of the collar where the bmp2/4 expression domain follows the submerged ectodermal track of the dorsal cord. (D) Section of a juvenile of a similar stage to (C). Dorsal is at the top of the panel. Note the submerged bmp domain, stained purple. (E) Dorsal view of bmp5/8 expression at day 3 of development with dorsal toward the viewer, and (F) in a hatching juvenile at day 5 of development. White arrowhead indicates the submerged domain in the collar (mesosome) similar to that of bmp2/4 in panel D. (G) Expression of bmp5/8 in the far posterior of a late juvenile at day 13 of development. White arrow indicates the position of the anus, before which the bmp5/8 domain stops. Posterior to the anus is the post anal tail. (H) Expression of tolloid/xolloid at day 3 of development. (I) Expression of bambi at the late gastrula stage. (J) Expression of crossveinless at the late gastrula stage and (K) at day 3 of development, with the dorsal midline oriented toward the viewer. (L) Tsg expression at day two of development. (M–P) Chordin expression from early to late developmental stages. (M) Expression at mid gastrula. (N) Sagittal section an embryo at day 2 of development just anterior to the telotroch; ventral is at the bottom of the panel. (O) Day 3 of development, a surface view of the lateral ectoderm, (P) and day 4 of development, in sagittal section. (Q) Admp and bmp2/4 double in situ hybridization at day 2 of development; brown is admp and blue is bmp2/4, and (R) a cross section of the same stage just anterior to the telotroch. (S) Admp expression at day 5 of development, and (T) surface view of an uncleared embryo, ventral midline toward the viewer.
Figure 3
Figure 3. Genes Expressed with Dorsal or Ventral Domains
These genes were chosen as candidate targets of Bmp activation or repression. All embryos are oriented as in Figure 2 with anterior to the top and left of each panel and dorsal in the top right of each panel, unless otherwise specified. The telotroch or ciliated band is marked by white arrowheads. Expression of dlx at (A) day 2 of development, just after gastrulation and (B) at day 3 of development. (C) Tbx 2/3 expression just after gastrulation, and (D) at day 3 of development. (E) Expression of hex at day 3 of development (F) Expression of nk2.3/2.5 at day 4. (G) Expression of olig on day 2, just after gastrulation, and (H) at day 3. (I) poxN expression at day 4 of development. (J) Pitx expression at day 2 of development, dorsal midline toward the viewer, a glancing optical section through the dorsal-most ectoderm. (K) Pitx expression at day 4 of development. Note the two domains of expression. (L) Netrin expression, a transverse section of a post-gastrula embryo at the level of the ciliated band. Note the broad ventral expression of netrin, and (M) the more narrow domain at day 3 of development. (N) Expression of lim3 at day 3 of development. (O) Expression of mnx at day 2 of development, and (P) at day 4. Note the ventral endodermal expression. (Q) Expression of mox (also called gax) at day 3 of development, and (R) a close up of the ventral domain at day 3, ventral midline toward the viewer, displaying the metasome and part of the mesosome. (S) Expression of sim at day 2 of development, and (T) at day 5.
Figure 4
Figure 4. Expression in S. kowalevskii of Orthologs of Chordate Genes Important in Dorsoventral Patterning of the Neural Tube
All embryos are shown as optical sections, and oriented in a similar manner as in Figure 2 with anterior to the top and left of each panel and dorsal in the top right of each panel, unless otherwise specified. (A) Hh expression in the apical tip of a day 3 embryo, and (B) at day 5 at the same location. (C) Endodermal expression of nk2–2 (also called nkx2.2) in the late gastrula, with a sharp delineation between presumptive proboscis mesoderm and definitive endoderm, and (D) at day 4, with expression continuing in the endoderm but down-regulated in the pouches of the forming gill slits (black arrowhead). (E) Expression of msx in the late gastrula, and (F) at day 3. Expression occurs exclusively in the ectoderm of the metasome and is down-regulated in the telotroch, the cilated band, as marked (white arrowhead).
Figure 5
Figure 5. Dorsalization of the S. kowalevskii Embryo by Application of Exogenous Bmp4 Protein
Exogenous Bmp4 protein was applied to the late blastula embryo (14 h post fertilization), and development was allowed to continue. All embryos are shown at a similar developmental stage, day 3 of development. (A) Side view of a control embryo cultured without Bmp4. The mouth is indicated by a black arrowhead on the ventral side. The normally developing endoderm shows a dorsal, anterior projection of the gut called the stomochord, indicated with a white arrowhead. One of the mesocoels is clearly visible on the dorsal midline, indicated by a yellow arrowhead. The endoderm is divided into two sections, the pharyngeal region in the anterior, divided from the posterior gut region by a posterior constriction shown by blue arrows. The first gills slit is indicated by a green arrowhead, just anterior to the gut division. (B) Embryos treated with 250ng/ml of Bmp 4, fixed at the same time as the control in panel A. The dorsoventral orientation is not possible to determine since they are cylindrically symmetric. Black arrowheads indicate thick condensations of mesenchyme around the anterior gut. (C) Embryos fixed at a similar development stage following a treatment with 500 ng/ml Bmp4 displaying a consistent phenotype between samples. Note the flattened anterior end and the thick connection of prosome and mesosome. (D) Expression of bmp2/4 following treatment with Bmp4 protein showing activation of endogenous expression throughout the ectoderm. (E) Stereomicrographs of uncleared embryos showing the expression of chordin in control embryos, with broad ventral expression, and (F) embryos following treatment with Bmp4 protein at 100 ng/ml. (G) Stereomicrographs of uncleared embryos showing the expression of elv in control embryos, with broad expression, but stronger at the midlines, and (H) embryos following treatment with Bmp4 protein at 100 ng/ml. Note the persistence of elv expression; it is not repressed by Bmp4. (I) Ubiquitous ectodermal expression of dlx following treatment with 100 ng/ml of Bmp4. (J) Expression of tbx2/3 expands throughout the ectoderm following treatment of the embryo with Bmp4 protein (250 ng/ml). White arrowheads indicate the position of the telotroch/ciliated band. (K) Expression of pitx expands from a spot to a ring around the base of the prosome in both the ectoderm and underlying mesenchyme, after Bmp4 protein treatment. (L) Control expression of hex at day 4 of development, and (M) following treatment with Bmp4 at 100ng/ml. (N) Pax1/9 expression expands from a dorsolateral spot to a circumferential ring in the endoderm following Bmp4 treatment at 100 ng/ml. (O) Like pax1/9, the nk2–3/2–5 domain expands from a short dorsal stripe to a ring in the endoderm, after Bmp4 treatment. (P) Expression of admp in the most anterior endoderm following treatment with 500 ng/ml of Bmp4. This is a residual spot (thus showing that the staining procedure has worked), whereas the entire ventral domains of ectoderm and endoderm have disappeared.
Figure 6
Figure 6. Ventralization of the S. kowalevskii Embryo following Injection of the Egg with Anti-bmp siRNAs
Eggs were injected immediately following fertilization with bmp siRNA and development was allowed to continue. (A) Low magnification micrograph of a group of embryos at the beginning of day 3 of development from injected eggs, showing the consistency of the phenotype. The dorsoventral orientation cannot be determined since the embryos are cylindrically symmetric. Note the deep indentation between the prosome and mesosome, and the short posterior end beyond the telotroch (ciliated band). (B) Optical section of an embryo at the same stage of development from an injected egg. (C) In the siRNA treated embryo, the expression of dlx continues in scattered cells in the anterior ectoderm of the prosome but disappears from the dorsal midline. (D) Expression of pitx at day 4 of development in a siRNA treated embryo, showing expanded expression throughout the majority of the metasome anterior to the telotroch, whereas the prosome dorsal spot is absent. The prosome has detached from the mesosome at this stage, as the mouth indentation encircles the embryo. The same developmental stage is shown in panel E and F. (E) Expression of admp expands strongly throughout the ectoderm of the detached prosome. (F) Low magnification micrograph of day 4 embryos showing expansion of expression of netrin from a ventral midline stripe to the entire ectoderm including that of the detached prosome. White arrowhead shows the position of the telotroch.
Figure 7
Figure 7. Summary of Inferred Evolutionary Changes of the Dorsoventral Axis in Deuterostome Evolution, with Emphasis on Hemichordates and Chordates
Mesoderm is shown in red, endoderm in yellow and ectoderm in blue (neural) and grey (epidermis). An ancestral secreted Bmp axis was involved in dorsoventral patterning of the three germ layers in the bilaterian ancestor. This ancestor we propose was characterized by a diffuse organization of its nervous system, shown by blue dots. A Bmp gradient was involved in dorsoventral patterning of all three germ layers. In the basal deuterostome and hemichordates the role of the Bmp gradient is conserved in general dorsoventral patterning. During chordate (and protostome) evolution, the existing Bmp/Chordin axis was co-opted for an additional developmental role in nervous system centralization.

Comment in

Similar articles

See all similar articles

Cited by 102 PubMed Central articles

See all "Cited by" articles

References

    1. De Robertis EM, Sasai Y. A common plan for dorsoventral patterning in Bilateria. Nature. 1996;380:37–40. - PubMed
    1. Lichtneckert R, Reichert H. Insights into the urbilaterian brain: Conserved genetic patterning mechanisms in insect and vertebrate brain development. Heredity. 2005;94:465–477. - PubMed
    1. Gerhart J, Lowe C, Kirschner M. Hemichordates and the origin of chordates. Curr Opin Genet Dev. 2005;15:461–467. - PubMed
    1. Finnerty JR, Pang K, Burton P, Paulson D, Martindale MQ. Origins of bilateral symmetry: Hox and dpp expression in a sea anemone. Science. 2004;304:1335–1337. - PubMed
    1. Martindale MQ, Finnerty JR, Henry JQ. The Radiata and the evolutionary origins of the bilaterian body plan. Mol Phylogenet Evol. 2002;24:358–365. - PubMed

Publication types

MeSH terms

LinkOut - more resources

Feedback