Embracing the diversity of model systems to deconstruct the basis of regeneration and tissue repair.

The eighth EMBO conference in the series 'The Molecular and Cellular Basis of Regeneration and Tissue Repair' took place in Barcelona (Spain) in September 2022. A total of 173 researchers from across the globe shared their latest advances in deciphering the molecular and cellular basis of wound healing, tissue repair and regeneration, as well as their implications for future clinical applications. The conference showcased an ever-expanding diversity of model organisms used to identify mechanisms that promote regeneration. Over 25 species were discussed, ranging from invertebrates to humans. Here, we provide an overview of the exciting topics presented at the conference, highlighting novel discoveries in regeneration and perspectives for regenerative medicine.

[The sea anemone Nematostella vectensis, an emerging model for biomedical research: Mechano-sensitivity, extreme regeneration and longevity]. / L'anémone de mer Nematostella vectensis - Un modèle émergent pour la recherche biomédicale : mécano-sensibilité, régénération et longévité.

Nematostella has fascinating features such as whole-body regeneration, the absence of signs of aging and importantly, the absence of age-related diseases. Easy to culture and spawn, this little sea anemone in spite of its "simple" aspect, displays interesting morphological characteristics similar to vertebrates and an unexpected similarity in gene content/genome organization. Importantly, the scientific community working on Nematostella is developing a variety of functional genomics tools that enable scientists to use this anemone in the field of regenerative medicine, longevity and mecano-sensory diseases. As a complementary research model to vertebrates, this marine invertebrate is emerging and promising to dig deeper into those fields of research in an integrative manner (entire organism) and provides new opportunities for scientists to lift specific barriers that can be encountered with other commonly used animal models.

TITLE: L'anémone de mer Nematostella vectensis - Un modèle émergent pour la recherche biomédicale : mécano-sensibilité, régénération et longévité. ABSTRACT: Nematostella, petite anémone de mer, possède de fascinantes propriétés, telles que la régénération du corps entier, l'absence de signes de vieillissement et d'affections liées à l'âge comme, par exemple, le développement de cancers. Elle se cultive aisément et se reproduit en laboratoire. Malgré son aspect « simple ¼, cet invertébré marin de l'embranchement des cnidaires partage avec les vertébrés des caractéristiques non seulement morphologiques, mais également génomiques. La communauté scientifique développe aujourd'hui une variété d'outils de génomique fonctionnelle permettant l'utilisation de cet animal de façon intégrative dans le domaine de la médecine régénérative, de la longévité et des maladies mécano-sensorielles. Son étude se présente comme particulièrement prometteuse pour faire progresser la connaissance dans ces différents domaines, offrant des possibilités expérimentales qui font défaut dans les modèles animaux classiques.

Experimental Tools to Study Regeneration in the Sea Anemone Nematostella vectensis.

Animal regeneration is a biological process leading to the reformation of injured or lost tissues/body parts. One of the most fascinating regenerative phenomena is the so-called whole-body regeneration, leading to the reformation of fully functional organisms within days after bisection. The sea anemone Nematostella vectensis is currently emerging as novel whole-body regeneration model. Here we describe the methods of inducing the regenerative process in this cnidarian as well as the fixation and staining protocols for morphological, molecular, and cellular analysis.

NvERTx: a gene expression database to compare embryogenesis and regeneration in the sea anemone Nematostella vectensis.

For over a century, researchers have been comparing embryogenesis and regeneration hoping that lessons learned from embryonic development will unlock hidden regenerative potential. This problem has historically been a difficult one to investigate because the best regenerative model systems are poor embryonic models and vice versa. Recently, however, there has been renewed interest in this question, as emerging models have allowed researchers to investigate these processes in the same organism. This interest has been further fueled by the advent of high-throughput transcriptomic analyses that provide virtual mountains of data. Here, we present Nematostella vectensis Embryogenesis and Regeneration Transcriptomics (NvERTx), a platform for comparing gene expression during embryogenesis and regeneration. NvERTx consists of close to 50 transcriptomic data sets spanning embryogenesis and regeneration in Nematostella These data were used to perform a robust de novo transcriptome assembly, with which users can search, conduct BLAST analyses, and plot the expression of multiple genes during these two developmental processes. The site is also home to the results of gene clustering analyses, to further mine the data and identify groups of co-expressed genes. The site can be accessed at http://nvertx.kahikai.org.

A bipolar role of the transcription factor ERG for cnidarian germ layer formation and apical domain patterning.

Germ layer formation and axial patterning are biological processes that are tightly linked during embryonic development of most metazoans. In addition to canonical WNT, it has been proposed that ERK-MAPK signaling is involved in specifying oral as well as aboral territories in cnidarians. However, the effector and the molecular mechanism underlying latter phenomenon is unknown. By screening for potential effectors of ERK-MAPK signaling in both domains, we identified a member of the ETS family of transcription factors, Nverg that is bi-polarily expressed prior to gastrulation. We further describe the crucial role of NvERG for gastrulation, endomesoderm as well as apical domain formation. The molecular characterization of the obtained NvERG knock-down phenotype using previously described as well as novel potential downstream targets, provides evidence that a single transcription factor, NvERG, simultaneously controls expression of two different sets of downstream targets, leading to two different embryonic gene regulatory networks (GRNs) in opposite poles of the developing embryo. We also highlight the molecular interaction of cWNT and MEK/ERK/ERG signaling that provides novel insight into the embryonic axial organization of Nematostella, and show a cWNT repressive role of MEK/ERK/ERG signaling in segregating the endomesoderm in two sub-domains, while a common input of both pathways is required for proper apical domain formation. Taking together, we build the first blueprint for a global cnidarian embryonic GRN that is the foundation for additional gene specific studies addressing the evolution of embryonic and larval development.

MAPK signaling is necessary for neurogenesis in Nematostella vectensis.

BACKGROUND: The nerve net of Nematostella is generated using a conserved cascade of neurogenic transcription factors. For example, NvashA, a homolog of the achaete-scute family of basic helix-loop-helix transcription factors, is necessary and sufficient to specify a subset of embryonic neurons. However, positive regulators required for the expression of neurogenic transcription factors remain poorly understood. RESULTS: We show that treatment with the MEK/MAPK inhibitor U0126 severely reduces the expression of known neurogenic genes, Nvath-like, NvsoxB(2), and NvashA, and known markers of differentiated neurons, suggesting that MAPK signaling is necessary for neural development. Interestingly, ectopic NvashA fails to rescue the expression of neural markers in U0126-treated animals. Double fluorescence in situ hybridization and transgenic analysis confirmed that NvashA targets represent both unique and overlapping populations of neurons. Finally, we used a genome-wide microarray to identify additional patterning genes downstream of MAPK that might contribute to neurogenesis. We identified 18 likely neural transcription factors, and surprisingly identified ~40 signaling genes and transcription factors that are expressed in either the aboral domain or animal pole that gives rise to the endomesoderm at late blastula stages. CONCLUSIONS: Together, our data suggest that MAPK is a key early regulator of neurogenesis, and that it is likely required at multiple steps. Initially, MAPK promotes neurogenesis by positively regulating expression of NvsoxB(2), Nvath-like, and NvashA. However, we also found that MAPK is necessary for the activity of the neurogenic transcription factor NvashA. Our forward molecular approach provided insight about the mechanisms of embryonic neurogenesis. For instance, NvashA suppression of Nvath-like suggests that inhibition of progenitor identity is an active process in newly born neurons, and we show that downstream targets of NvashA reflect multiple neural subtypes rather than a uniform neural fate. Lastly, analysis of the MAPK targets in the early embryo suggests that MAPK signaling is critical not only to neurogenesis, but also endomesoderm formation and aboral patterning.

Regulative capacity for eye formation by first quartet micromeres of the polychaete Capitella teleta.

The stereotypic cleavage pattern shared by spiralian embryos provides unique opportunities to compare mechanisms of cell fate specification of homologous blastomeres, and can give insights into how changes in fate may have influenced the evolution of novel structures and morphological diversity. The potential of cells to undergo regulation and the timing of cell fate specification were investigated during early development in the polychaete annelid, Capitella teleta. Targeted laser deletions of the first quartet micromeres were performed, with a focus on the eye-forming cells 1a and 1c. Most of the larvae resulting from deletion of the 1a or 1c micromeres lack both the pigment cell and sensory cell of the eye as predicted by the C. teleta fate map. In a minority of cases, however, both left and right larval eye spots develop, suggesting that other blastomeres within the embryo regulate for loss of these cells. Deletion of the 1a and 1c derivatives, 1a(1) or 1c(1), also largely result in larvae with one pigment spot, although there are larvae with two eye spots, suggesting that the ability to regulate for loss of an eye-generating cell persists for an additional cell cycle. Cell deletion in conjunction with intracellular labeling indicates that all four quadrants retain the ability to generate eyes, including those that normally do not. Deletion of all four first quartet micromeres provides evidence that only the first quartet micromeres have eye-forming potential. Additionally, in contrast to the right side of the head where larval and adult eye sensory cells are derived from the same cell (1c), on the left side, the larval and adult eye sensory cells are generated by different embryonic lineages. We hypothesize that cell-cell interactions and cell position are important for regulative ability in Capitella. To our knowledge, this is one of the first detailed deletion studies of the first quartet micromeres and the first convincing example of regulation in polychaetes, which are often thought to be non-regulative in nature.

Characterization of Morphological and Cellular Events Underlying Oral Regeneration in the Sea Anemone, Nematostella vectensis.

Cnidarians, the extant sister group to bilateria, are well known for their impressive regenerative capacity. The sea anemone Nematostella vectensis is a well-established system for the study of development and evolution that is receiving increased attention for its regenerative capacity. Nematostella is able to regrow missing body parts within five to six days after its bisection, yet studies describing the morphological, cellular, and molecular events underlying this process are sparse and very heterogeneous in their experimental approaches. In this study, we lay down the basic framework to study oral regeneration in Nematostella vectensis. Using various imaging and staining techniques we characterize in detail the morphological, cellular, and global molecular events that define specific landmarks of this process. Furthermore, we describe in vivo assays to evaluate wound healing success and the initiation of pharynx reformation. Using our described landmarks for regeneration and in vivo assays, we analyze the effects of perturbing either transcription or cellular proliferation on the regenerative process. Interestingly, neither one of these experimental perturbations has major effects on wound closure, although they slightly delay or partially block it. We further show that while the inhibition of transcription blocks regeneration in a very early step, inhibiting cellular proliferation only affects later events such as pharynx reformation and tentacle elongation.

Nodal signaling is required for mesodermal and ventral but not for dorsal fates in the indirect developing hemichordate, Ptychodera flava.

Nodal signaling plays crucial roles in vertebrate developmental processes such as endoderm and mesoderm formation, and axial patterning events along the anteroposterior, dorsoventral and left-right axes. In echinoderms, Nodal plays an essential role in the establishment of the dorsoventral axis and left-right asymmetry, but not in endoderm or mesoderm induction. In protostomes, Nodal signaling appears to be involved only in establishing left-right asymmetry. Hence, it is hypothesized that Nodal signaling has been co-opted to pattern the dorsoventral axis of deuterostomes and for endoderm, mesoderm formation as well as anteroposterior patterning in chordates. Hemichordata, together with echinoderms, represent the sister taxon to chordates. In this study, we analyze the role of Nodal signaling in the indirect developing hemichordate Ptychodera flava. In particular, we show that during gastrulation nodal transcripts are detected in a ring of cells at the vegetal pole that gives rise to endomesoderm and in the ventral ectoderm at later stages of development. Inhibition of Nodal function disrupts dorsoventral fates and also blocks formation of the larval mesoderm. Interestingly, molecular analysis reveals that only mesodermal, apical and ventral gene expression is affected while the dorsal side appears to be patterned correctly. Taken together, this study suggests that the co-option of Nodal signaling in mesoderm formation and potentially in anteroposterior patterning has occurred prior to the emergence of chordates and that Nodal signaling on the ventral side is uncoupled from BMP signaling on the dorsal side, representing a major difference from the molecular mechanisms of dorsoventral patterning events in echinoderms.

An organizing activity is required for head patterning and cell fate specification in the polychaete annelid Capitella teleta: new insights into cell-cell signaling in Lophotrochozoa.

Many lophotrochozoans (i.e., molluscs, annelids, nemerteans, and polyclad flatworms) display a well-conserved early developmental program called spiral cleavage that contrasts with the high diversity of adult body forms present in this group. Due to this stereotypical development, each cell can be uniquely identified and its lineage history known following intracellular injection of lineage tracers. Cell deletion experiments performed mainly in molluscs have demonstrated that one or two cells associated with the endomesodermal lineage represent an embryonic organizer of subsequent development and are causally involved in cell fate and body patterning. Utilizing the published fate map of the spiral-cleaving annelid Capitella teleta, we used infrared laser cell deletions to dissect the role of individual cells on the patterning of the larval body. Thirteen uniquely identifiable individual blastomeres and two double cell combination deletions were studied to assess larval phenotypes by scoring multiple morphological structures and cell type-specific molecular markers differentially expressed along the antero-posterior and dorso-ventral axes. Surprisingly, our results show that in C. teleta, the cellular identity of the "organizing cell" and the timing of the organizing activity are different from that of other spiralians. retain-->In C. teleta, the ectodermal primary somatoblast, 2d, is the key cell responsible for organizing activity during early embryonic development, and is necessary for bilateral symmetry and dorso-ventral axis organization of the head as well as neural, foregut and mesoderm tissue formation. Furthermore, we show that the ERK/MAPK signaling pathway does not appear to be involved in organizing activity in retain-->C. teleta. This contrasts with data from molluscs and the molecular mechanism suggested for another polychaete, Hydroides elegans, highlighting likely molecular level variation among spiralian embryos. These results reinforce the idea that an embryonic organizing activity is present across spiralians. Our data also emphasize the developmental variation within lophotrochozoans, and may ultimately provide insight into the role of developmental processes in the evolution of diverse body forms in metazoans.