Hunger and satiety: Neuronal dynamics in Hydra behavior.

Hormonal and neuronal inputs to the brain control how much animals eat. The origins of this behavior were unclear, but in this issue of Cell Reports, Giez et al.1 describe specific neurons inhibiting feeding in evolutionary ancient animals without brain.

A new look at the architecture and dynamics of the Hydra nerve net.

The Hydra nervous system is the paradigm of a 'simple nerve net'. Nerve cells in Hydra, as in many cnidarian polyps, are organized in a nerve net extending throughout the body column. This nerve net is required for control of spontaneous behavior: elimination of nerve cells leads to polyps that do not move and are incapable of capturing and ingesting prey (Campbell, 1976). We have re-examined the structure of the Hydra nerve net by immunostaining fixed polyps with a novel antibody that stains all nerve cells in Hydra. Confocal imaging shows that there are two distinct nerve nets, one in the ectoderm and one in the endoderm, with the unexpected absence of nerve cells in the endoderm of the tentacles. The nerve nets in the ectoderm and endoderm do not contact each other. High-resolution TEM (transmission electron microscopy) and serial block face SEM (scanning electron microscopy) show that the nerve nets consist of bundles of parallel overlapping neurites. Results from transgenic lines show that neurite bundles include different neural circuits and hence that neurites in bundles require circuit-specific recognition. Nerve cell-specific innexins indicate that gap junctions can provide this specificity. The occurrence of bundles of neurites supports a model for continuous growth and differentiation of the nerve net by lateral addition of new nerve cells to the existing net. This model was confirmed by tracking newly differentiated nerve cells.

The origin and evolution of Wnt signalling.

The Wnt signal transduction pathway has essential roles in the formation of the primary body axis during development, cellular differentiation and tissue homeostasis. This animal-specific pathway has been studied extensively in contexts ranging from developmental biology to medicine for more than 40 years. Despite its physiological importance, an understanding of the evolutionary origin and primary function of Wnt signalling has begun to emerge only recently. Recent studies on very basal metazoan species have shown high levels of conservation of components of both canonical and non-canonical Wnt signalling pathways. Furthermore, some pathway proteins have been described also in non-animal species, suggesting that recruitment and functional adaptation of these factors has occurred in metazoans. In this Review, we summarize the current state of research regarding the evolutionary origin of Wnt signalling, its ancestral function and the characteristics of the primal Wnt ligand, with emphasis on the importance of genomic studies in various pre-metazoan and basal metazoan species.

The significance of Ethel Browne's research on Hydra for the organizer concept.

This article focuses on the roots of the organizer concept, which was developed by Hans Spemann during his studies of early embryonic development in amphibians. The fundamental properties of this axis-inducing signaling center have been elucidated through pioneering molecular research by Eddy De Robertis' laboratory and other researchers. Evolutionary comparisons have disclosed the presence of this signaling center, involving the interaction of Wnt and TGF-beta signaling pathways, existed not only in vertebrates but also in basal Metazoa such as Cnidaria. - Notably, even prior to the groundbreaking experiments conducted by Hilde Mangold and Hans Spemann, Ethel Browne conducted similar transplantation experiments on Hydra polyps. They were performed under the guidance of Thomas H Morgan and in the laboratory of Edmund B Wilson. Howard Lenhoff was the first to draw connections between Ethel Browne's transplantation experiments and those of Spemann and Mangold, igniting a vivid debate on the precedence of the organizer concept and its recognition in Nobel Prize considerations. This review critically compares the experiments conducted by Spemann and Mangold with those preceding their seminal work, concluding that the organizer concept clearly builds upon earlier research aimed at understanding developmental gradients, such as in the simple model Hydra. However, these approaches were not pursued further by Morgan, who shifted his focus towards unraveling the genetic control of development in flies, an approach that ultimately revealed the molecular identity of the Spemann organizer in vertebrates.

Reversible Host-Guest Crosslinks in Supramolecular Hydrogels for On-Demand Mechanical Stimulation of Human Mesenchymal Stem Cells.

Stem cells are regulated not only by biochemical signals but also by biophysical properties of extracellular matrix (ECM). The ECM is constantly monitored and remodeled because the fate of stem cells can be misdirected when the mechanical interaction between cells and ECM is imbalanced. A well-defined ECM model for bone marrow-derived human mesenchymal stem cells (hMSCs) based on supramolecular hydrogels containing reversible host-guest crosslinks is fabricated. The stiffness (Young's modulus E) of the hydrogels can be switched reversibly by altering the concentration of non-cytotoxic, free guest molecules dissolved in the culture medium. Fine-adjustment of substrate stiffness enables the authors to determine the critical stiffness level E* at which hMSCs turn the mechano-sensory machinery on or off. Next, the substrate stiffness across E* is switched and the dynamic adaptation characteristics such as morphology, traction force, and YAP/TAZ signaling of hMSCs are monitored. These data demonstrate the instantaneous switching of traction force, which is followed by YAP/TAZ signaling and morphological adaptation. Periodical switching of the substrate stiffness across E* proves that frequent applications of mechanical stimuli drastically suppress hMSC proliferation. Mechanical stimulation across E* level using dynamic hydrogels is a promising strategy for the on-demand control of hMSC transcription and proliferation.

Evolution: Neuronal control of an archaic mouth.

Cnidarians (corals, hydras, jellyfish, sea anemones) are prey-devouring creatures with a simple nervous system, the function of which is largely unknown. A new study on the freshwater polyp Hydra has now uncovered the neuronal circuits that control its feeding behavior.

A predatory gastrula leads to symbiosis-independent settlement in Aiptasia.

The planula larvae of the sea anemone Aiptasia have so far not been reported to complete their life cycle by undergoing metamorphosis into adult forms. This has been a major obstacle in their use as a model for coral-dinoflagellate endosymbiosis. Here, we show that Aiptasia larvae actively feed on crustacean nauplii, displaying a preference for live prey. This feeding behavior relies on functional stinging cells, indicative of complex neuronal control. Regular feeding leads to significant size increase, morphological changes, and efficient settlement around 14 d postfertilization. Surprisingly, the presence of dinoflagellate endosymbionts does not affect larval growth or settlement dynamics but is crucial for sexual reproduction. Our findings finally close Aiptasia's life cycle and highlight the functional nature of its larvae, as in Haeckel's Gastrea postulate, yet reveal its active carnivory, thus contributing to our understanding of early metazoan evolution.

Wnt/ß-catenin signaling induces axial elasticity patterns of Hydra extracellular matrix.

The extracellular matrix (ECM) plays crucial roles in animal development and diseases. Here, we report that Wnt/ß-catenin signaling induces the ECM remodeling during Hydra axis formation. We determined the micro- and nanoscopic arrangement of fibrillar type I collagen along Hydra's body axis using high-resolution microscopy and X-ray scattering. Elasticity mapping of the ECM ex vivo revealed distinctive elasticity patterns along the body axis. A proteomic analysis of the ECM showed that these elasticity patterns correlate with a gradient-like distribution of metalloproteases along the body axis. Activation of the Wnt/ß-catenin pathway in wild-type and transgenic animals alters these patterns toward low ECM elasticity patterns. This suggests a mechanism whereby high protease activity under control of Wnt/ß-catenin signaling causes remodeling and softening of the ECM. This Wnt-dependent spatiotemporal coordination of biochemical and biomechanical cues in ECM formation was likely a central evolutionary innovation for animal tissue morphogenesis.

The Hydra stem cell system - Revisited.

Cnidarians are >600 million years old and are considered the sister group of Bilateria based on numerous molecular phylogenetic studies. Apart from Hydra, the genomes of all major clades of Cnidaria have been uncovered (e.g. Aurelia, Clytia, Nematostella and Acropora) and they reveal a remarkable completeness of the metazoan genomic toolbox. Of particular interest is Hydra, a model system of aging research, regenerative biology, and stem cell biology. With the knowledge gained from scRNA research, it is now possible to characterize the expression profiles of all cell types with great precision. In functional studies, our picture of the Hydra stem cell biology has changed, and we are in the process of obtaining a clear picture of the homeostasis and properties of the different stem cell populations. Even though Hydra is often compared to plant systems, the new data on germline and regeneration, but also on the dynamics and plasticity of the nervous system, show that Hydra with its simple body plan represents in a nutshell the prototype of an animal with stem cell lineages, whose properties correspond in many ways to Bilateria. This review provides an overview of the four stem cell lineages, the two epithelial lineages that constitute the ectoderm and the endoderm, as well as the multipotent somatic interstitial lineage (MPSC) and the germline stem cell lineage (GSC), also known as the interstitial cells of Hydra.

From injury to patterning-MAPKs and Wnt signaling in Hydra.

Hydra has a regenerative capacity that is not limited to individual organs but encompasses the entire body. Various global and integrative genome, transcriptome and proteome approaches have shown that many of the signaling pathways and transcription factors present in vertebrates are already present in Cnidaria, the sister group of Bilateria, and are also activated in regeneration. It is now possible to investigate one of the central questions of regeneration biology, i.e., how does the patterning system become activated by the injury signals that initiate regeneration. This review will present the current data obtained in Hydra and draw parallels with regeneration in Bilateria. Important findings of this global analysis are that the Wnt signaling pathway has a dual function in the regeneration process. In the early phase Wnt is activated generically and in a second phase of pattern formation it is activated in a position specific manner. Thus, Wnt signaling is part of the generic injury response, in which mitogen-activated protein kinases (MAPKs) are initially activated via calcium and reactive oxygen species (ROS). The MAPKs, p38, c-Jun N-terminal kinases (JNKs) and extracellular signal-regulated kinases (ERK) are essential for Wnt activation in Hydra head and foot regenerates. Furthermore, the antagonism between the ERK signaling pathway and stress-induced MAPKs results in a balanced induction of apoptosis and mitosis. However, the early Wnt genes are activated by MAPK signaling rather than apoptosis. Early Wnt gene activity is differentially integrated with a stable, ß-Catenin-based gradient along the primary body axis maintaining axial polarity and activating further Wnts in the regenerating head. Because MAPKs and Wnts are highly evolutionarily conserved, we hypothesize that this mechanism is also present in vertebrates but may be activated to different degrees at the level of early Wnt gene integration.

Non-muscle myosin II drives critical steps of nematocyst morphogenesis.

Nematocysts are generated by secretion of proteins into a post-Golgi compartment. They consist of a capsule that elongates into a long tube, which is coiled inside the capsule matrix and expelled during its nano-second discharge deployed for prey capture. The driving force for discharge is an extreme osmotic pressure of 150 bar. The complex processes of tube elongation and invagination under these biomechanical constraints have so far been elusive. Here, we show that a non-muscle myosin II homolog (HyNMII) is essential for nematocyst formation in Hydra. In early nematocysts, HyNMII assembles to a collar around the neck of the protruding tube. HyNMII then facilitates tube outgrowth by compressing it along the longitudinal axis as evidenced by inhibitor treatment and genetic knockdown. In addition, live imaging of a NOWA::NOWA-GFP transgenic line, which re-defined NOWA as a tube component facilitating invagination, allowed us to analyze the impact of HyNMII on tube maturation.

Injury-induced MAPK activation triggers body axis formation in Hydra by default Wnt signaling.

Hydra's almost unlimited regenerative potential is based on Wnt signaling, but so far it is unknown how the injury stimulus is transmitted to discrete patterning fates in head and foot regenerates. We previously identified mitogen-activated protein kinases (MAPKs) among the earliest injury response molecules in Hydra head regeneration. Here, we show that three MAPKs-p38, c-Jun N-terminal kinases (JNKs), and extracellular signal-regulated kinases (ERKs)-are essential to initiate regeneration in Hydra, independent of the wound position. Their activation occurs in response to any injury and requires calcium and reactive oxygen species (ROS) signaling. Phosphorylated MAPKs hereby exhibit cross talk with mutual antagonism between the ERK pathway and stress-induced MAPKs, orchestrating a balance between cell survival and apoptosis. Importantly, Wnt3 and Wnt9/10c, which are induced by MAPK signaling, can partially rescue regeneration in tissues treated with MAPK inhibitors. Also, foot regenerates can be reverted to form head tissue by a pharmacological increase of ß-catenin signaling or the application of recombinant Wnts. We propose a model in which a ß-catenin-based stable gradient of head-forming capacity along the primary body axis, by differentially integrating an indiscriminate injury response, determines the fate of the regenerating tissue. Hereby, Wnt signaling acquires sustained activation in the head regenerate, while it is transient in the presumptive foot tissue. Given the high level of evolutionary conservation of MAPKs and Wnts, we assume that this mechanism is deeply embedded in our genome.

The role of cnidarian developmental biology in unraveling axis formation and Wnt signaling.

Cnidarians are fascinating creatures at the base of metazoan evolution possessing an almost unlimited regeneration capacity that has attracted the interest of researchers, from Abraham Trembley's discovery of regeneration to the present. They share a simple body plan and a high morphogenetic plasticity that has led to a broad spectrum of life cycles. With molecular genomics it became unequivocally clear that Cnidaria are the sister group of the Bilateria and how similar their molecular toolkit is to that of more complex animals. This has renewed interest in these simple animals, which have had an important role in the establishment of fundamental concepts for developmental biologists from the beginning. This review focuses on our current understanding of signaling centers (organizers) and morphogenetic gradients in cnidarians and how they relate to the emergence of the bilaterian body axes. The data are largely based on the cnidarian models Hydra and Nematostella and are supported by new studies on forms with a complete cnidarian life cycle, such as the medusozoans Aurelia and Clytia. Molecular studies on cnidarian development have revealed the existence of an ancient Wnt signaling center at the site of gastrulation, which was instrumental for the formation of a primary body axis and can be traced back to the common ancestor of bilaterian and non-bilaterian animals. New molecular data also suggest that the molecular vectors for the dorso-ventral and left-right body axis in bilaterians, Bmp and Nodal signaling, respectively, were already present but had different fates in the two clades. The close link of developmental processes in bilaterians and cnidarians but also their distinct differences make cnidarians an indispensable model for tackling fundamental questions in developmental biology from patterning, regeneration and other recent molecular approaches to theoretical concepts.

Two plus one is almost three: a fast approximation for multi-view deconvolution.

Multi-view deconvolution is a powerful image-processing tool for light sheet fluorescence microscopy, providing isotropic resolution and enhancing the image content. However, performing these calculations on large datasets is computationally demanding and time-consuming even on high-end workstations. Especially in long-time measurements on developing animals, huge amounts of image data are acquired. To keep them manageable, redundancies should be removed right after image acquisition. To this end, we report a fast approximation to three-dimensional multi-view deconvolution, denoted 2D+1D multi-view deconvolution, which is able to keep up with the data flow. It first operates on the two dimensions perpendicular and subsequently on the one parallel to the rotation axis, exploiting the rotational symmetry of the point spread function along the rotation axis. We validated our algorithm and evaluated it quantitatively against two-dimensional and three-dimensional multi-view deconvolution using simulated and real image data. 2D+1D multi-view deconvolution takes similar computation time but performs markedly better than the two-dimensional approximation only. Therefore, it will be most useful for image processing in time-critical applications, where the full 3D multi-view deconvolution cannot keep up with the data flow.

The Wnt-specific astacin proteinase HAS-7 restricts head organizer formation in Hydra.

BACKGROUND: The Hydra head organizer acts as a signaling center that initiates and maintains the primary body axis in steady state polyps and during budding or regeneration. Wnt/beta-Catenin signaling functions as a primary cue controlling this process, but how Wnt ligand activity is locally restricted at the protein level is poorly understood. Here we report a proteomic analysis of Hydra head tissue leading to the identification of an astacin family proteinase as a Wnt processing factor. RESULTS: Hydra astacin-7 (HAS-7) is expressed from gland cells as an apical-distal gradient in the body column, peaking close beneath the tentacle zone. HAS-7 siRNA knockdown abrogates HyWnt3 proteolysis in the head tissue and induces a robust double axis phenotype, which is rescued by simultaneous HyWnt3 knockdown. Accordingly, double axes are also observed in conditions of increased Wnt activity as in transgenic actin::HyWnt3 and HyDkk1/2/4 siRNA treated animals. HyWnt3-induced double axes in Xenopus embryos could be rescued by coinjection of HAS-7 mRNA. Mathematical modelling combined with experimental promotor analysis indicate an indirect regulation of HAS-7 by beta-Catenin, expanding the classical Turing-type activator-inhibitor model. CONCLUSIONS: We show the astacin family protease HAS-7 maintains a single head organizer through proteolysis of HyWnt3. Our data suggest a negative regulatory function of Wnt processing astacin proteinases in the global patterning of the oral-aboral axis in Hydra.

Expansion of a single transposable element family is associated with genome-size increase and radiation in the genus Hydra.

Transposable elements are one of the major contributors to genome-size differences in metazoans. Despite this, relatively little is known about the evolutionary patterns of element expansions and the element families involved. Here we report a broad genomic sampling within the genus Hydra, a freshwater cnidarian at the focal point of diverse research in regeneration, symbiosis, biogeography, and aging. We find that the genome of Hydra is the result of an expansion event involving long interspersed nuclear elements and in particular a single family of the chicken repeat 1 (CR1) class. This expansion is unique to a subgroup of the genus Hydra, the brown hydras, and is absent in the green hydra, which has a repeat landscape similar to that of other cnidarians. These features of the genome make Hydra attractive for studies of transposon-driven genome expansions and speciation.

A combined strategy of neuropeptide prediction and tandem mass spectrometry identifies evolutionarily conserved ancient neuropeptides in the sea anemone Nematostella vectensis.

Neuropeptides are a class of bioactive peptides shown to be involved in various physiological processes, including metabolism, development, and reproduction. Although neuropeptide candidates have been predicted from genomic and transcriptomic data, comprehensive characterization of neuropeptide repertoires remains a challenge owing to their small size and variable sequences. De novo prediction of neuropeptides from genome or transcriptome data is difficult and usually only efficient for those peptides that have identified orthologs in other animal species. Recent peptidomics technology has enabled systematic structural identification of neuropeptides by using the combination of liquid chromatography and tandem mass spectrometry. However, reliable identification of naturally occurring peptides using a conventional tandem mass spectrometry approach, scanning spectra against a protein database, remains difficult because a large search space must be scanned due to the absence of a cleavage enzyme specification. We developed a pipeline consisting of in silico prediction of candidate neuropeptides followed by peptide-spectrum matching. This approach enables highly sensitive and reliable neuropeptide identification, as the search space for peptide-spectrum matching is highly reduced. Nematostella vectensis is a basal eumetazoan with one of the most ancient nervous systems. We scanned the Nematostella protein database for sequences displaying structural hallmarks typical of eumetazoan neuropeptide precursors, including amino- and carboxyterminal motifs and associated modifications. Peptide-spectrum matching was performed against a dataset of peptides that are cleaved in silico from these putative peptide precursors. The dozens of newly identified neuropeptides display structural similarities to bilaterian neuropeptides including tachykinin, myoinhibitory peptide, and neuromedin-U/pyrokinin, suggesting these neuropeptides occurred in the eumetazoan ancestor of all animal species.

Membrane capacitance recordings resolve dynamics and complexity of receptor-mediated endocytosis in Wnt signalling.

Receptor-mediated endocytosis is an essential process in signalling pathways for activation of intracellular signalling cascades. One example is the Wnt signalling pathway that seems to depend on endocytosis of the ligand-receptor complex for initiation of Wnt signal transduction. To date, the roles of different endocytic pathways in Wnt signalling, molecular players and the kinetics of the process remain unclear. Here, we monitored endocytosis in Wnt3a and Wnt5a-mediated signalling with membrane capacitance recordings of HEK293 cells. Our measurements revealed a swift and substantial increase in the number of endocytic vesicles. Extracellular Wnt ligands specifically triggered endocytotic activity, which started immediately upon ligand binding and ceased within a period of ten minutes. By using specific inhibitors, we were able to separate Wnt-induced endocytosis into two independent pathways. We demonstrate that canonical Wnt3a is taken up mainly by clathrin-independent endocytosis whereas noncanonical Wnt5a is exclusively regulated via clathrin-mediated endocytosis. Our findings show that membrane capacitance recordings allow the resolution of complex cellular processes in plasma membrane signalling pathways in great detail.

Regionalized nervous system in Hydra and the mechanism of its development.

The last common ancestor of Bilateria and Cnidaria is considered to develop a nervous system over 500 million years ago. Despite the long course of evolution, many of the neuron-related genes, which are active in Bilateria, are also found in the cnidarian Hydra. Thus, Hydra is a good model to study the putative primitive nervous system in the last common ancestor that had the great potential to evolve to a more advanced one. Regionalization of the nervous system is one of the advanced features of bilaterian nervous system. Although a regionalized nervous system is already known to be present in Hydra, its developmental mechanisms are poorly understood. In this study we show how it is formed and maintained, focusing on the neuropeptide Hym-176 gene and its paralogs. First, we demonstrate that four axially localized neuron subsets that express different combination of the neuropeptide Hym-176 gene and its paralogs cover almost an entire body, forming a regionalized nervous system in Hydra. Second, we show that positional information governed by the Wnt signaling pathway plays a key role in determining the regional specificity of the neuron subsets as is the case in bilaterians. Finally, we demonstrated two basic mechanisms, regionally restricted new differentiation and phenotypic conversion, both of which are in part conserved in bilaterians, are involved in maintaining boundaries between the neuron subsets. Therefore, this study is the first comprehensive analysis of the anatomy and developmental regulation of the divergently evolved and axially regionalized peptidergic nervous system in Hydra, implicating an ancestral origin of neural regionalization.