Isolation of a substance activating foot formation in hydra

We have developed an assay for a substance from hydra that accelerates foot regeneration in the animal. This substance is specific for the foot as evidenced by the following findings: (1) It is present in the animal as a steep gradient descending from foot to head, paralleling the foot-forming potential of the tissue (2) It does not accelerate head regeneration, nor do the head factors of hydra discovered by Schaller (1973) and Berking (1977) accelerate foot regeneration. We propose that the foot-activating substance is a morphogen responsible for foot formation in hydra. The foot activator can be extracted from hydra tissue with methanol and separated from other known morphogens of hydra by gel filtration and ion-exchange chromatography. A substance with similar biological and physicochemical properties can be isolated from sea anemones.     

Ultraviolet irradiation initiates ectopic foot formation in regenerating hydra and promotes budding

We have studied the effects of ultraviolet-C (UVC) and Ultraviolet-B (UVB) on growth and pattern formation inPelmatohydra oligactis. UVC brings about a significant increase in budding in intact hydra while UVB does not exhibit such an effect. Excessive budding could be a response for survival at wavelengths that damage biological tissues. If the head or base piece of a bisected hydra is irradiated and recombined with the unirradiated missing part, regeneration proceeds normally indicating that exposure of a body part with either an intact head or foot to UVC does not influence pattern formation. Most significantly, in the middle piece, but not in the head or the base piece of a trisected hydra, UVC leads to initiation of ectopic feet formation in almost one third of the cases. Thus, UV irradiation interferes with pattern formation in regenerating hydra, possibly by changing positional values, and promotes budding in intact hydra. This is the first report on induction of ectopic feet formation by UV in regenerating hydra and opens up the possibility of using UV irradiation as a tool to understand pattern formation in the enigmatic hydra     

The foot formation stimulating peptide pedibin is also involved in patterning of the head in hydra

Avian neural crest cells migrate on precise pathways to their target areas where they form a wide variety of cellular derivatives, including neurons, glia, pigment cells and skeletal components. In one portion of their pathway, trunk neural crest cells navigate in the somitic mesoderm in a segmental fashion, invading the rostral, while avoiding the caudal, half-sclerotome. This pattern of cell migration, imposed by the somitic mesoderm, contributes to the metameric organization of the peripheral nervous system, including the sensory and sympathetic ganglia. At hindbrain levels, neural crest cells also travel from the neural tube in a segmental manner via three migratory streams of cells that lie adjacent to even-numbered rhombomeres. In this case, the adjacent mesoderm does not possess an obvious segmental organization, compared to the somitic mesoderm at trunk levels. Thus, the mechanisms by which the embryo controls segmentally-organized cell migrations have been a fascinating topic over the past several years. Here, I discuss findings from classical and recent studies that have delineated several of the tissue, cellular and molecular elements that contribute to the segmental organization of neural crest migration, primarily in the avian embryo. One common theme is that neural crest cells are prohibited from entering particular territories in the embryo due to the expression of inhibitory factors. However, permissive, migration-promoting factors may also play a key role in coordinating neural crest migration.     

Properties of the foot inhibitor from hydra

Summary A substance was isolated from crude extracts of hydra that inhibits foot regeneration. This substance, the foot inhibitor, has a molecular weight of 500 daltons. It is a hydrophilic molecule, slightly basic in character and it has no peptide bonds. The pruified substance acts specifically and at concentrations lower than 10^–7 M. At this low concentration only foot and not head regeneration is inhibited. Hydra are sensitive to purified foot inhibitor between the second and eight hour after initiation of foot regeneration by cutting. In normal animals the foot inhibitor is most likely produced by nerve cells. A substance with similar biological and physico-chemical properties is found in other coelenterates.     

Receptor-based models with hysteresis for pattern formation in hydra

In this paper, we propose a new receptor-based model for pattern formation and regulation in a fresh-water polyp, namely hydra. The model is defined in the form of a system of reaction-diffusion equations with zero-flux boundary conditions coupled with a system of ordinary differential equations. The production of diffusible biochemical molecules has a hysteretic dependence on the density of these molecules and is modeled by additional ordinary differential equations. We study the hysteresis-driven mechanism of pattern formation and we demonstrate the advantages and constraints of its ability to explain different aspects of pattern formation and regulation in hydra. The properties of the model demonstrate a range of stationary and oscillatory spatially heterogeneous patterns, arising from multiple spatially homogeneous steady states and switches in the production rates.     

The head and the foot inhibitor from hydra are not dowex artefacts

Summary In a recent publication in this journal (Berking 1983) it was claimed (1) that the head inhibitor we isolated from hydra is a Dowex artefact, (2) that a separate foot inhibitor does not exist in hydra and (3) that the only inhibitor that has so far been isolated from hydra is one which inhibits head and foot regeneration equally well. These statements are incorrect and require a response. In the following, I would like to summarise our evidence that the inhibitors isolated from hydra, including Berking's inhibitor, have different specificities for head and foot regeneration. In addition, I would like to show that none of our substances are Dowex artefacts.     

Tissue healing and septate desmosome formation in hydra

Tissue healing was studied in hydra tissue grafts by means of light and electron microscopy. Healing is begun by the gastrodermis: subsequently the epidermis fuses and the mesoglea is repaired. Epidermis fusion is first brought about by long processes from the basal portions of the epithelial cells bridging the wound gap and adhering to opposing cells. Irregular septate desmosomes form early in this process and continuously become more neatly organized. Concommitant with the healing process at the graft site, neighboring cells are also rearranging, their septate desmosomes undergoing transient disorganizations. We conclude that the organization of septate junctions is dynamic, and may be undergoing a balanced but continuous, steady state turnover. During the healing process the forces acting on the desmosomes, and other aspects of the cells' architecture, are not balanced, and the junctions grow and become more highly organized.     

Pattern formation in hydra tissue without developmental gradients

Ring-shaped pieces of hydra tissue were excised from a specified position on a body column of 20-30 polyps and grafted together in tandem like a chain of beads. A "tandem graft" prepared in this way has the same basic tissue organization and same tube-like morphology as a normal hydra body column, but lacks the head, foot, and developmental gradients ordinarily present. Three major types of structures were formed along the length of the tandem graft: heads, buds, and feet. The relative number of these structures produced was strongly affected by the origin of the tissue used to prepare the tandem graft. Evidence was obtained which suggests that tissue originally located outside of the budding zone in intact hydra has a strong latent capacity to form a bud, and that the level of this capacity forms a gradient from the budding zone toward the hypostome. Evidence was also obtained which is consistent with the view that the head and foot forming mechanisms cross-react positively, increasing the chances for these two structures to be formed next to each other on a tandem graft.     

Action of foot activator on growth and differentiation of cells in hydra

Foot activator is a small peptide found in hydra and specifically activates foot formation. I present a method for the further purification of foot activator by high-pressure liquid chromatography. The morphogenetically active fractions were assayed for their effect at the cellular level. Foot activator acts as a mitogen by pushing epithelial and interstitial cells, which are arrested in G2, into mitosis. In the presence of foot activator, epithelial stem cells are stimulated to differentiate into foot mucus cells, and interstitial nerve precursor cells differentiate into mature nerve cells. The interaction of foot activator with head activator in the development of hydra is discussed.     

Tumour-promoting phorbol esters rapidly inhibit bud formation in hydra

Summary The effects of tumour promoters and carcinogens on bud formation were investigated in an attempt to clarify the primary process of bud formation in hydra. Treatment with 1.0ng/ml 12-O-tetradecanoylphorbol-13-acetate (TPA), phorbol-12,13-didecanoate (PDD) or mezerein added immediately after feeding rapidly and completely inhibited the formation of new buds in Hydra japonica. Treatment with TPA 3–6 h after feeding also suppressed bud formation 24 h later, but suppressed buds appeared 48 h later. Buds suppressed by TPA also formed in the presence of a diluted homogenate of hydra and during starvation. Carcinogens, such as benzo(a)pyrene and 20-methylcholanthrene, did not have an inhibitory effect on bud formation within 2 days. The tumour promoters and carcinogens used in this experiment did not inhibit the regeneration of tentacles. These results indicate that tumour-promoting phorbol esters, but not carcinogens, rapidly suppress the process by which the formation of buds is initiated by hydra, and the effects of these esters depend on the timing of treatment after feeding.     

The interstitial cells control the sexual phenotype of heterosexual chimeras of hydra

The three stem cell populations in hydra, the epithelial cells of the ectoderm and endoderm, which make up the body of the hydra, and the interstitial cells, which give rise to nerve cells, nematocytes, and gametes, were tested for their effects on determining the sexual phenotype of individuals. This was done by creating epithelial hydra, which are devoid of interstitial cells and their derivatives, of one sexual type and repopulating them with interstitial cells from individuals of the other sexual type. The resulting heterosexual chimeras were found in all cases to display the same sexual phenotype as that of the interstitial cell donor, indicating this cell type is responsible for the sex of the animal. The epithelial tissue had no influence in determining which gamete type was produced.     

Cell-cell interactions and the control of sex determination in hydra

Sexual phenotype in gonochoristic species of hydra (species having separate males and females) is ultimately determined by cells committed to the sperm pathway. Little is known about the genetic basis for determining sex during embryogenesis, but the sex ratio of offspring is generally 1 : 1, implying a genetic component. Sexual phenotype of adult hydra is labile and sex-reversal in both directions can be induced by experimental manipulations involving addition or loss of the sperm lineage. Loss of the sperm lineage is induced by high temperature, causing males to switch to females. Introduction of the sperm lineage into females by grafting causes sex-reversal to male. These studies provide support for a model of sex determination based on cell-cell interactions.     

A model for budding in hydra: pattern formation in concentric rings

Current models of pattern formation in Hydra propose head-and foot-specific morphogens to control the development of the body ends and along the body length axis. In addition, these morphogens are proposed to control a cellular parameter (positional value, source density) which changes gradually along the axis. This gradient determines the tissue polarity and the regional capacity to form a head and a foot, respectively, in transplantation experiments. The current models are very successful in explaining regeneration and transplantation experiments. However, some results obtained render problems, in particular budding, the asexual way of reproduction is not understood. Here an alternative model is presented to overcome these problems. A primary system of interactions controls the positional values. At certain positional values secondary systems become active which initiate the local formation of e.g. mouth, tentacles, and basal disc. (i) A system of autocatalysis and lateral inhibition is suggested to exist as proposed by Gierer and Meinhardt (Kybernetik 12 (1972) 30). (ii) The activator is neither a head nor a foot activator but rather causes an increase of the positional value. (iii) On the other hand, a generation of the activator leads to its loss from cells and therewith to a (local) decrease of the positional value. (iv) An inhibitor is proposed to exist which antagonizes an increase of the positional value. External conditions like the gradient of positional values in the surroundings and interactions with other sites of morphogen production decide whether at a certain site of activator generation the positional value will increase (head formation), decrease (foot formation) or increase in the centre and decrease in the periphery thereby forming concentric rings (bud formation). Computer-simulation experiments show basic features of budding, regeneration and transplantation.     

HyAlx, an aristaless-related gene, is involved in tentacle formation in hydra

Developmental gradients are known to play important roles in axial patterning in hydra. Current efforts are directed toward elucidating the molecular basis of these gradients. We report the isolation and characterization of HyAlx, an aristaless-related gene in hydra. The expression patterns of the gene in adult hydra, as well as during bud formation, head regeneration and the formation of ectopic head structures along the body column, indicate the gene plays a role in the specification of tissue for tentacle formation. The use of RNAi provides more direct evidence for this conclusion. The different patterns of HyAlx expression during head regeneration and bud formation also provide support for a recent version of a reaction-diffusion model for axial patterning in hydra.