Strabismus-mediated primary archenteron invagination is uncoupled from Wnt/β-catenin-dependent endoderm cell fate specification in Nematostella vectensis (Anthozoa, Cnidaria): Implications for the evolution of gastrulation

<p>Abstract</p> <p>Background</p> <p>Gastrulation is a uniquely metazoan character, and its genesis was arguably the key step that enabled the remarkable diversification within this clade. The process of gastrulation involves two tightly coupled events during embryogenesis of most metazoans. Morphogenesis produces a distinct internal epithelial layer in the embryo, and this epithelium becomes segregated as an endoderm/endomesodermal germ layer through the activation of a specific gene regulatory program. The developmental mechanisms that induced archenteron formation and led to the segregation of germ layers during metazoan evolution are unknown. But an increased understanding of development in early diverging taxa at the base of the metazoan tree may provide insights into the origins of these developmental mechanisms.</p> <p>Results</p> <p>In the anthozoan cnidarian <it>Nematostella vectensis</it>, initial archenteron formation begins with bottle cell-induced buckling of the blastula epithelium at the animal pole. Here, we show that bottle cell formation and initial gut invagination in <it>Nematostella </it>requires NvStrabismus (NvStbm), a maternally-expressed core component of the Wnt/Planar Cell Polarity (PCP) pathway. The NvStbm protein is localized to the animal pole of the zygote, remains asymmetrically expressed through the cleavage stages, and becomes restricted to the apical side of invaginating bottle cells at the blastopore. Antisense morpholino-mediated NvStbm-knockdown blocks bottle cell formation and initial archenteron invagination, but it has no effect on Wnt/ß-catenin signaling-mediated endoderm cell fate specification. Conversely, selectively blocking Wnt/ß-catenin signaling inhibits endoderm cell fate specification but does not affect bottle cell formation and initial archenteron invagination.</p> <p>Conclusions</p> <p>Our results demonstrate that Wnt/PCP-mediated initial archenteron invagination can be uncoupled from Wnt/ß-catenin-mediated endoderm cell fate specification in <it>Nematostella</it>, and provides evidence that these two processes could have evolved independently during metazoan evolution. We propose a two-step model for the evolution of an archenteron and the evolution of endodermal germ layer segregation. Asymmetric accumulation and activation of Wnt/PCP components at the animal pole of the last common ancestor to the eumetazoa may have induced the cell shape changes that led to the initial formation of an archenteron. Activation of Wnt/ß-catenin signaling at the animal pole may have led to the activation of a gene regulatory network that specified an endodermal cell fate in the archenteron.</p

Evolutionary transition between invertebrates and vertebrates via methylation reprogramming in embryogenesis

© The Author(s), 2019. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Xu, X., Li, G., Li, C., Zhang, J., Wang, Q., Simmons, D. K., Chen, X., Wijesena, N., Zhu, W., Wang, Z., Wang, Z., Ju, B., Ci, W., Lu, X., Yu, D., Wang, Q., Aluru, N., Oliveri, P., Zhang, Y. E., Martindale, M. Q., & Liu, J. Evolutionary transition between invertebrates and vertebrates via methylation reprogramming in embryogenesis. National Science Review, 6(5), (2019):993-1003, doi:10.1093/nsr/nwz064.Major evolutionary transitions are enigmas, and the most notable enigma is between invertebrates and vertebrates, with numerous spectacular innovations. To search for the molecular connections involved, we asked whether global epigenetic changes may offer a clue by surveying the inheritance and reprogramming of parental DNA methylation across metazoans. We focused on gametes and early embryos, where the methylomes are known to evolve divergently between fish and mammals. Here, we find that methylome reprogramming during embryogenesis occurs neither in pre-bilaterians such as cnidarians nor in protostomes such as insects, but clearly presents in deuterostomes such as echinoderms and invertebrate chordates, and then becomes more evident in vertebrates. Functional association analysis suggests that DNA methylation reprogramming is associated with development, reproduction and adaptive immunity for vertebrates, but not for invertebrates. Interestingly, the single HOX cluster of invertebrates maintains unmethylated status in all stages examined. In contrast, the multiple HOX clusters show dramatic dynamics of DNA methylation during vertebrate embryogenesis. Notably, the methylation dynamics of HOX clusters are associated with their spatiotemporal expression in mammals. Our study reveals that DNA methylation reprogramming has evolved dramatically during animal evolution, especially after the evolutionary transitions from invertebrates to vertebrates, and then to mammals.This work was supported by the National Key Research and Development Program of China (2018YFC1003303), the Strategic Priority Research Program of the CAS (XDB13040200), the National Natural Science Foundation of China (91519306, 31425015), the Youth Innovation Promotion Association of the CAS and the Key Research Program of Frontier Sciences, CAS (QYZDY-SSW-SMC016)

Wnt Signaling in the Cnidarian Nematostella vectensis: Insighits into the Evolution of Gastrulation

The genesis of gastrulation was arguably a key evolutionary innovation that enabled metazoan diversification. The developmental mechanisms that induced archenteron formation and endodermal cell fate specification during gastrulation are unknown, but one crucial step was likely the co-option of a localized molecular asymmetry that was present in ancient embryos. One ancient polarity present in most metazoan eggs is the animal vegetal (AV) or primary egg axis, which is established by the asymmetric localization of maternal factors such as RNA, proteins and organelles during oogenesis. In bilaterians, which account for a majority of animal taxa, the AV axis predicts the axial properties of the embryo and the adult with the animal pole giving rise to the anterior of the organism, and the vegetal pole giving rise to the endomesoderm and the site of gastrulation. In contrast, in early diverging non-bilaterian taxa such as ctenophores and cnidarians, the animal pole gives rise to the endoderm and the site of gastrulation. These and other observations have led to the hypothesis that endoderm specification and gastrulation evolved at the animal pole, and moreover, that the mechanisms that regulate these processes were moved down to the vegetal pole in the bilaterian lineage. This idea is supported at the molecular level by the observation that endoderm specification in both bilaterians and non-bilaterians is regulated by nuclear beta-catenin signaling indicating a role for the Wnt signaling pathway in the evolution of gastrulation. In the cnidarian Nematostella vectensis, the Dishevelled (NvDsh) protein, a critical component of Wnt signaling, is enriched at the animal pole of eggs and embryos and is required for Wnt/beta-catenin signaling mediated endodermal cell fate specification but not for primary archenteron invagination. Primary archenteron invagination in Nematostella is mediated by the localized activation of Wnt/PCP signaling at the animal pole. The mechanisms coordinating Wnt signaling dependent endoderm cell fate specification and primary archenteron invagination at the animal pole in Nematostella embryos are not known, but elucidation of these mechanisms may provide critical insight into the evolution of gastrulation. There is increasing evidence to suggest that a highly conserved group of core proteins functioning in the Wnt/PCP signaling pathway play a critical role in regulating gastrulation through the asymmetric localization of key components of signaling cascades in both vertebrates and invertebrates. For my dissertation research, I focused on the role of core Wnt/PCP proteins in establishing the embryonic polarity that leads to the asymmetric activation of Wnt signaling in the blastomeres at the animal pole in Nematostella. In Nematostella, the core PCP genes NvStrabismus (NvStbm) and NvFlamingo (NvFmi) are expressed maternally and are localized to the animal pole from the egg stage throughout early development. The Frizzled homologs NvFrizzled1 (NvFz1), NvFrizzled4 (NvFz4) and NvFrizzed5 (NvFz5) are expressed maternally in dynamic expression patterns during early development. NvFz10 is expressed in the early gastrula stage at the animal pole and is restricted to the invaginating cells of the presumptive endoderm. NvPrickle (NvPk) is asymmetrically expressed at the blastula stage and at the late gastrula stage it is expressed in the pharynx and on one side of the embryo in the presumptive ectoderm. Inhibition of NvFz1 function during early development of Nematostella blocked endodermal cell fate specification but not primary archenteron invagination. In contrast, loss of NvFz10 function blocked primary archenteron invagination without affecting endodermal cell fate specification. This experimental uncoupling of Wnt/PCP signaling mediated initial archenteron invagination from Wnt/beta-catenin mediated endoderm specification in Nematostella provides further evidence for the independent evolution of these two processes during early metazoan evolution. The localized expression of NvFmi to the animal pole of eggs and embryos indicated a possible role for this core Wnt/PCP protein in coordinating the activity of the two Wnt pathways at the animal pole. This idea was tested by disrupting NvFmi function using morpholinos and dominant-negative approaches. Downregulation of NvFmi function blocked both Wnt/PCP signaling mediated archenteron invagination and Wnt/beta-catenin signaling mediated endoderm cell fate specification, while overexpressing the cytoplasmic carboxy terminus of the NvFmi protein selectively disrupted endodermal gene expression. These observations indicates that NvFmi regulates both Wnt pathways and that it may function as a scaffold to coordinate both Wnt/?-catenin and Wnt/PCP signaling to drive primary archenteron invagination and endoderm cell fate specification in Nematostella through the asymmetric localization of Wnt pathway components to the animal pole. Overall, this study has provided experimental evidence as to how different branches of Wnt signaling mediate primary archenteron invagination and cell fate specification through NvFz10 and NvFz1 respectively during gastrulation in Nematostella and possible mechanisms of coordination of these two processes in time and space through the function of NvFmi.</p

Distinct Frizzled receptors independently mediate endomesoderm specification and primary archenteron invagination during gastrulation in Nematostella

Endomesodermal cell fate specification and archenteron formation during gastrulation are tightly linked developmental processes in most metazoans. However, studies have shown that in the anthozoan cnidarian Nematostella vectensis, Wnt/β-catenin (cWnt) signalling-mediated endomesodermal cell fate specification can be experimentally uncoupled from Wnt/Planar Cell Polarity (PCP) signalling-mediated primary archenteron invagination. The upstream signalling mechanisms regulating cWnt signalling-dependent endomesoderm cell fate specification and Wnt/PCP signalling-mediated primary archenteron invagination in Nematostella embryos are not well understood. By screening for potential upstream mediators of cWnt and Wnt/PCP signalling, we identified two Nematostella Frizzled homologs that are expressed early in development. NvFzd1 is expressed maternally and in a broad pattern during early development while NvFzd10 is zygotically expressed at the animal pole in blastula stage embryos and is restricted to the invaginating cells of the presumptive endomesoderm. Molecular and morphological characterization of NvFzd1 and NvFzd10 knock-down phenotypes provide evidence for distinct regulatory roles for the two receptors in endomesoderm cell fate specification and primary archenteron invagination. These results provide further experimental evidence for the independent regulation of endomesodermal cell fate specification and primary archenteron invagination during gastrulation in Nematostella. Moreover, these results provide additional support for the previously proposed two-step model for the independent evolution of cWnt-mediated cell fate specification and Wnt/PCP-mediated primary archenteron invagination. [Display omitted] •Endomesoderm specification and gastrulation are tightly linked events in Bilateria.•These processes can be experimentally uncoupled in the cnidarian N. vectensis.•Nematostella primary gut invagination can occur without endomesoderm specification.•NvFzd1 regulates endomesoderm specification and NvFzd10 regulates gut invagination.•These results suggest that the two processes arose independently during evolution