The Hydra genome: insights, puzzles and opportunities for developmental biologists

The sequencing of a Hydra genome marked the beginning of a new era in the use of Hydra as a developmental model. Analysis of the genome sequence has led to a number of interesting findings, has required revisiting of previous work, and most importantly presents new opportunities for understanding the developmental biology of Hydra. This review will de-scribe the history of the Hydra genome project, a selection of results from it that are relevant to developmental biologists, and some future research opportunities provided by Hydra genomics.     

GFP expression in Hydra: lessons from the particle gun

The cnidarian Hydra is an important model organism to study pattern formation and tem cell differentiation. In the past, however, it has been difficult to study gene function in Hydra because the animals have hot been accessible to gene transfection studies, we have now developed a method to transiently express GFP-tagged proteins in Hydra using a green fluorescent protein (GFP) expression plasmid under the control of the Hydra actin promoter and a particle gun to introduce it into Hydra cell nuclei. We achieve strong transient GFP expression in a small but reproducible number of epithelial and interstitial cells. Implications for the use of this method to carry out single cell assays with GFP-tagged Hydra proteins are discussed.     

Germline stem cells and sex determination in Hydra

The sex of germline stem cells (GSCs) in Hydra is determined in a cell-autonomous manner. In gonochoristic species like Hydra magnipapillata or H. oligactis, where the sexes are separate, male polyps have sperm-restricted stem cells (SpSCs), while females have egg-restricted stem cells (EgSCs). These GSCs self-renew in a polyp, and are usually transmitted to a new bud from a parental polyp during asexual reproduction. But if these GSCs are lost during subsequent budding or regeneration events, new ones are generated from multipotent stem cells (MPSCs). MPSCs are the somatic stem cells in Hydra that ordinarily differentiate into nerve cells, nematocytes (stinging cells in cnidarians), and gland cells. By means of such a backup system, sexual reproduction is guaranteed for every polyp. Interestingly, Hydra polyps occasionally undergo sex-reversal. This implies that each polyp can produce either type of GSCs, i.e. Hydra are genetically hermaphroditic. Nevertheless a polyp possesses only one type of GSCs at a time. We propose a plausible model for sex-reversal in Hydra. We also discuss so-called germline specific genes, which are expressed in both GSCs and MPSCs, and some future plans to investigate Hydra GSCs.     

The Synaptonemal Complex of Basal Metazoan Hydra:More Similarities to Vertebrate than Invertebrate Meiosis Model Organisms

The synaptonemal complex （SC） is an evolutionarily well-conserved structure that mediates chromosome synapsis during prophase of the first meiotic division. Although its structure is conserved, the characterized protein components in the current metazoan meiosis model systems （Drosophila melanogaster, Caenorhabditis elegans, and Mus musculus） show no sequence homology, challenging the question of a single evolutionary origin of the SC. However, our recent studies revealed the monophyletic origin of the mammalian SC protein components. Many of them being ancient in Metazoa and already present in the cnidarian Hydra. Remarkably, a comparison between different model systems disclosed a great similarity between the SC components of Hydra and mammals while the proteins of the ecdysozoan systems （D. rnelanogaster and C. elegans） differ significantly. In this review, we introduce the basal-branching metazoan species Hydra as a potential novel invertebrate model system for meiosis research and particularly for the investigation of SC evolution, function and assembly. Also, available methods for SC research in Hydra are summarized.     

The Hydra Model System

The freshwater Hydra polyp emerged as a model system in 1741 when Abraham Trembley not only discovered its amazing regenerative potential, but also demonstrated that experimental manipulations pave the way to research in biology. Since then, Hydra flourished as a potent and fruitful model system to help answer questions linked to cell and developmental biology, as such as the setting up of an organizer to regenerate a complex missing structure, the establishment and maintainance of polarity in a multicellular organism, the development of mathematical models to explain the robust developmental rules observed in this animal, the maintainance of stemness and multipotency in a highly dynamic environment, the plasticity of differentiated cells, to name but a few. However the Hydra model system is not restricted to cell and developmental biology; during the past 270 years it has also been heavily used to investigate the relationships between Hydra and its environment, opening new horizons concerning neurophysiology, innate immunity, ecosystems, ecotoxicology, symbiosis...     

Hydra, a model system for environmental studies

Hydra have been extensively used for studying the teratogenic and toxic potential of numerous toxins throughout the years and are more recently growing in popularity to assess the impacts of environmental pollutants. Hydra are an appropriate bioindicator species for use in environmental assessment owing to their easily measurable physical (morphology), biochemical (xenobiotic biotransformation; oxidative stress), behavioural (feeding) and reproductive (sexual and asexual) endpoints. Hydra also possess an unparalleled ability to regenerate, allowing the assessment of teratogenic compounds and the impact of contaminants on stem cells. Importantly, Hydra are ubiquitous throughout freshwater environments and relatively easy to culture making them appropriate for use in small scale bioassay systems. Hydra have been used to assess the environmental impacts of numerous environmental pollutants including metals, organic toxicants (including pharmaceuticals and endocrine disrupting compounds), nanomaterials and industrial and municipal effluents. They have been found to be among the most sensitive animals tested for metals and certain effluents, comparing favourably with more standardised toxicity tests. Despite their lack of use in formalised monitoring programmes, Hydra have been extensively used and are regarded as a model organism in aquatic toxicology.     

Developmental signaling in Hydra: what does it take to build a "simple" animal?

Developmental processes in multicellular animals depend on an array of signal transduction pathways. Studies of model organisms have identified a number of such pathways and dissected them in detail. However, these model organisms are all bilaterians. Investigations of the roles of signal transduction pathways in the early-diverging metazoan Hydra have revealed that a number of the well-known developmental signaling pathways were already in place in the last common ancestor of Hydra and bilaterians. In addition to these shared pathways, it appears that developmental processes in Hydra make use of pathways involving a variety of peptides. Such pathways have not yet been identified as developmental regulators in more recently diverged animals. In this review I will summarize work to date on developmental signaling pathways in Hydra and discuss the future directions in which such work will need to proceed to realize the potential that lies in this simple animal.     

Value of the Hydra model system for studying symbiosis

Green Hydra is used as a classical example for explaining symbiosis in schools as well as an excellent research model. Indeed the cosmopolitan green Hydra (Hydra viridissima) provides a potent experimental framework to investigate the symbiotic relationships between a complex eumetazoan organism and a unicellular photoautotrophic green algae named Chlorella. Chlorella populates a single somatic cell type, the gastrodermal myoepithelial cells (also named digestive cells) and the oocyte at the time of sexual reproduction. This symbiotic relationship is stable, well-determined and provides biological advantages to the algal symbionts, but also to green Hydra over the related non-symbiotic Hydra i.e. brown hydra. These advantages likely result from the bidirectional flow of metabolites between the host and the symbiont. Moreover genetic flow through horizontal gene transfer might also participate in the establishment of these selective advantages. However, these relationships between the host and the symbionts may be more complex. Thus, Jolley and Smith showed that the reproductive rate of the algae increases dramatically outside of Hydra cells, although this endosymbiont isolation is debated. Recently it became possible to keep different species of endosymbionts isolated from green Hydra in stable and permanent cultures and compare them to free-living Chlorella species. Future studies testing metabolic relationships and genetic flow should help elucidate the mechanisms that support the maintenance of symbiosis in a eumetazoan species.     

The Hydra model - a model for what?

The introductory personal remarks refer to my motivations for choosing research projects, and for moving from physics to molecular biology and then to development, with Hydra as a model system. Historically, Trembley's discovery of Hydra regeneration in 1744 was the beginning of developmental biology as we understand it, with passionate debates about preformation versus de novo generation, mechanisms versus organisms. In fact, seemingly conflicting bottom-up and top-down concepts are both required in combination to understand development. In modern terms, this means analysing the molecules involved, as well as searching for physical principles underlying development within systems of molecules, cells and tissues. During the last decade, molecular biology has provided surprising and impressive evidence that the same types of molecules and molecular systems are involved in pattern formation in a wide range of organisms, including coelenterates like Hydra, and thus appear to have been "invented" early in evolution. Likewise, the features of certain systems, especially those of developmental regulation, are found in many different organisms. This includes the generation of spatial structures by the interplay of self-enhancing activation and "lateral" inhibitory effects of wider range, which is a main topic of my essay. Hydra regeneration is a particularly clear model for the formation of defined patterns within initially near-uniform tissues. In conclusion, this essay emphasizes the analysis of development in terms of physical laws, including the application of mathematics, and insists that Hydra was, and will continue to be, a rewarding model for understanding general features of embryogenesis and regeneration.     

What Hydra can teach us about chemical ecology -how a simple, soft organism survives in a hostile aqueous environment

Hydra and its fellow cnidarians - sea anemones, corals and jellyfish - are simple, mostly sessile animals that depend on bioactive chemicals for survival. In this review, we briefly describe what is known about the chemical armament of Hydra, and detail future research directions where Hydra can help illuminate major questions in chemical ecology, pharmacology, developmental biology and evolution. Focusing on two groups of putative toxins from Hydra - phospholipase A2s and proteins containing ShK and zinc metalloprotease domains, we ask: how do different venom components act together during prey paralysis? How is a venom arsenal created and how does it evolve? How is the chemical arsenal delivered to its target? To what extent does a chemical and biotic coupling exist between an organism and its environment? We propose a model whereby in Hydra and other cnidarians, bioactive compounds are secreted both as localized point sources (nematocyte discharges) and across extensive body surfaces, likely combining to create complex "chemical landscapes". We speculate that these cnidarian-derived chemical landscapes may affect the surrounding community on scales from microns to, in the case of coral reefs, hundreds of kilometers.     

Hydra, a fruitful model system for 270 years

The discovery of Hydra regeneration by Abraham Trembley in 1744 promoted much scientific curiosity thanks to his clever design of experimental strategies away from the natural environment. Since then, this little freshwater cnidarian polyp flourished as a potent and fruitful model system. Here, we review some general biological questions that benefitted from Hydra research, such as the nature of embryogenesis, neurogenesis, induction by organizers, sex reversal, symbiosis, aging, feeding behavior, light regulation, multipotency of somatic stem cells, temperature-induced cell death, neuronal transdifferentiation, to cite only a few. To understand how phenotypes arise, theoricists also chose Hydra to model patterning and morphogenetic events, providing helpful concepts such as reaction-diffusion, positional information, and autocatalysis combined with lateral inhibition. Indeed, throughout these past 270 years, scientists used transplantation and grafting experiments, together with tissue, cell and molecular labelings, as well as biochemical procedures, in order to establish the solid foundations of cell and developmental biology. Nowadays, thanks to transgenic, genomic and proteomic tools, Hydra remains a promising model for these fields, but also for addressing novel questions such as evolutionary mechanisms, maintenance of dynamic homeostasis, regulation of stemness, functions of autophagy, cell death, stress response, innate immunity, bioactive compounds in ecosystems, ecotoxicant sensing and science communication.     

Ancient signals: peptides and the interpretation of positional information in ancestral metazoans

Understanding the 'tool kit' that builds the most fundamental aspects of animal complexity requires data from the basal animals. Among the earliest diverging animal phyla are the Cnidaria which are the first in having a defined body plan including an axis, a nervous system and a tissue layer construction. Here I revise our understanding of patterning mechanism in cnidarians with special emphasis on the nature of positional signals in Hydra as perhaps the best studied model organism within this phylum. I show that (i) peptides play a major role as positional signals and in cell-cell communication; (ii) that intracellular signalling pathways in Hydra leading to activation of target genes are shared with all multicellular animals; (iii) that homeobox genes translate the positional signals; and (iv) that the signals are integrated by a complex genetic regulatory machinery that includes both novel cis regulatory elements as well as taxon specific target genes. On the basis of these results I present a model for the regulatory interactions required for axis formation in Hydra.     

Cloning and characterisation of PKB and PRK homologs from Hydra and the evolution of the protein kinase family

Two new serine/threonine protein kinases have been cloned from Hydra cDNA. The first of these kinases belongs to the PKB/Akt family. It is expressed ubiquitously in Hydra at a relatively low level but is upregulated during head regeneration. The second kinase is a member of the PRK/PKN family. It is ubiquitously expressed in Hydra tissue, albeit at a higher level than PKB. Construction of a phylogenetic tree including the Hydra PRK and PKB kinases and two PKC homologs previously cloned by Hassel and comparing them with members of the PKC, PKB and PRK families from porifera, Dictyostelium,yeast, Drosophila, Caenorhabditis and humans provide support for a simple model for the evolution of these kinase families. An ancestral precursor which contained a pleckstrin homology domain in its N-terminus and a C-terminal kinase domain gave rise to PKB in Dictyostelium. From this ancestor the PKB/PRK and PKC families evolved. The pleckstrin homology domain was lost in the PKC and PRK families and kept in the PKB family. PKB homologs have now been found in a variety of multicellular animals with Hydra being the phylogenetically earliest representative. Members of the PRK/PKC family, on the other hand, are also present in fungi. The precursor for these kinases must have contained N-terminal regulatory domains that were retained in fungal PRKs but subsequently partitioned between kinases of the PKC and PRK groups in metazoans.     

Hymyc1 downregulation promotes stem cell proliferation in Hydra vulgaris

Hydra is a unique model for studying the mechanisms underlying stem cell biology. The activity of the three stem cell lineages structuring its body constantly replenishes mature cells lost due to normal tissue turnover. By a poorly understood mechanism, stem cells are maintained through self-renewal while concomitantly producing differentiated progeny. In vertebrates, one of many genes that participate in regulating stem cell homeostasis is the protooncogene c-myc, which has been recently identified also in Hydra, and found expressed in the interstitial stem cell lineage. In the present paper, by developing a novel strategy of RNA interference-mediated gene silencing (RNAi) based on an enhanced uptake of small interfering RNAi (siRNA), we provide molecular and biological evidence for an unexpected function of the Hydra myc gene (Hymyc1) in the homeostasis of the interstitial stem cell lineage. We found that Hymyc1 inhibition impairs the balance between stem cell self renewal/differentiation, as shown by the accumulation of stem cell intermediate and terminal differentiation products in genetically interfered animals. The identical phenotype induced by the 10058-F4 inhibitor, a disruptor of c-Myc/Max dimerization, demonstrates the specificity of the RNAi approach. We show the kinetic and the reversible feature of Hymyc1 RNAi, together with the effects displayed on regenerating animals. Our results show the involvement of Hymyc1 in the control of interstitial stem cell dynamics, provide new clues to decipher the molecular control of the cell and tissue plasticity in Hydra, and also provide further insights into the complex myc network in higher organisms. The ability of Hydra cells to uptake double stranded RNA and to trigger a RNAi response lays the foundations of a comprehensive analysis of the RNAi response in Hydra allowing us to track back in the evolution and the origin of this process.     

Components, structure, biogenesis and function of the Hydra extracellular matrix in regeneration, pattern formation and cell differentiation

The body wall of Hydra is organized as an epithelial bilayer (ectoderm and endoderm) with an intervening extracellular matrix (ECM), termed mesoglea by early biologists. Morphological studies have determined that Hydra ECM is composed of two basal lamina layers positioned at the base of each epithelial layer with an intervening interstitial matrix. Molecular and biochemical analyses of Hydra ECM have established that it contains components similar to those seen in more complicated vertebrate species. These components include such macromolecules as laminin, type IV collagen, and various fibrillar collagens. These components are synthesized in a complicated manner involving cross-talk between the epithelial bilayer. Any perturbation to ECM biogenesis leads to a blockage in Hydra morphogenesis. Blockage in ECM/cell interactions in the adult polyp also leads to problems in epithelial transdifferentiation processes. In terms of biophysical parameters, Hydra ECM is highly flexible; a property that facilitates continuous movements along the organism's longitudinal and radial axis. This is in contrast to the more rigid matrices often found in vertebrates. The flexible nature of Hydra ECM can in part now be explained by the unique structure of the organism's type IV collagen and fibrillar collagens. This review will focus on Hydra ECM in regard to: 1) its general structure, 2) its molecular composition, 3) the biophysical basis for the flexible nature of Hydra's ECM, 4) the relationship of the biogenesis of Hydra ECM to regeneration of body form, and 5) the functional role of Hydra ECM during pattern formation and cell differentiation.     

水螅摄食中的特殊行为

取经中性红活染后的杆吻虫喂水螅,仔细观察了水螅的摄食行为.结果表明,水螅垂唇端部能够相互识别出同类;水螅的垂唇与胃区能协力把杆吻虫吞入胃腔,触手经常随食物进入胃腔;吞食时经常出现头部内翻或外翻;个体间及其成体与芽体间常出现争夺食物的现象,当垂唇端部互不相触时,体型较大的个体常常把较小个体或芽体连同食物一起吞食;垂唇内胚层腺细胞对食物有消化作用,对同类无伤害;水螅的神经系统已有初步的整合功能.     

First identification and localization of a visual pigment in Hydra (Cnidaria, Hydrozoa)

The cnidarian Hydra does not possess identified photoreceptive structures or specialized cells for light detection; nevertheless, it shows a marked photosensitivity. So far no evidence has been previously reported about the localization of the proteins involved in the photoresponse. We used polyclonal antibodies and immunofluorescence microscopy on whole-mount Hydra to identify a putative rhodopsin-like protein. Our results show an immunoreactivity in the ectodermal layer of Hydra, which corresponds in position to the nervous epidermal sensory cells. These data provide the first identification of a rhodopsin-like protein in a phylogenetically old invertebrate and give a new insight into the Hydra photoreceptive response.     

Molecular Cloning of a Preprohormone from Hydra magnipapillata Containing Multiple Copies of Hydra-LWamide (Leu-Trp-NH2) Neuropeptides: Evidence for Processing at Ser and Asn Residues

Abstract: The simple, freshwater polyp Hydra is often used as a model to study development in cnidarians. Recently, a neuropeptide, 2, has been isolated from sea anemones that induces metamorphosis in a hydroid planula larva to become a polyp. Here, we have cloned a preprohormone from Hydra magnipapillata containing 11 (eight different) immature neuropeptide sequences that are structurally related to the metamorphosis-inducing neuropeptide from sea anemones. During the final phase of our cloning experiments, another research team independently isolated and sequenced five of the neuropeptides originally found on the preprohormone. Comparison of these mature neuropeptide structures with the immature neuropeptide sequences on the preprohormone shows that most immature neuropeptide sequences are preceded by Ser or Asn residues, indicating that these residues must be novel processing sites. Thus, the structure of the Hydra prepro-hormone confirms our earlier findings that cnidarian pre-prohormones contain unusual or novel processing sites. Nearly all neuropeptide copies located on the Hydra preprohormone will give rise to mature neuropeptides with a C-terminal Gly-Leu-Trp-NH₂ sequence (the most frequent one being Gly-Pro-Pro-Pro-Gly-Leu-Trp-NH₂; Hydra-LWamide I; three copies). Based on their structural similarities with the metamorphosis-inducing neuropeptide from sea anemones, the mature peptides derived from the Hydra-LWamide preprohormone are potential candidates for being developmentally active neurohormones in Hydra .