Chemical composition of spines and tests of sea urchins

The skeleton of spines and tests of the species of sea urchins Strongylocentrotus intermedius, Mesocentrotus nudus, Scaphechinus mirabilis, and Echinocardium cordatum from the Sea of Japan is composed of a spongy stereom, consisting of calcite with a high content of magnesium. It was found that the tests and spines of the skeletons of sea urchins are composed of calcium–organic composite materials inlaid with other metals: Mg, Fe, Zn, and Rb. In the four species of sea urchins studied, the strength and other mechanical properties of the tests and spines differ and depend on the chemical composition and structural organization of their components. It was shown that the content of volatile substances correlates with their fragility or elasticity. It is revealed that the chemical composition of the tests of two species of the spherical sea urchins S. intermedius and M. nudus indicates significant differences between these two species of sea urchins.     

The molecular evolution of sperm bindin in six species of sea urchins (Echinoida: Strongylocentrotidae)

The acrosomal protein bindin attaches sperm to eggs during sea urchin fertilization. Complementary to ongoing functional biochemical studies, I take a comparative approach to explore the molecular evolution of bindin in a group of closely related free-spawning echinoid species. Two alleles of the mature bindin gene were sequenced for each of six species in the sea urchin family Strongylocentrotidae. The nucleotide sequences diverged by at least 1% per Myr at both silent and replacement sites. Two short sections flanking the conserved block show an excess of nonsynonymous substitutions. Each is homologous to a region that had been identified as a target of selection in other sea urchin comparisons. A large proportion of the bindin-coding sequence consists of a highly variable repeat region. Bindin sequences, even including the large intron, could not resolve the branching order among five of the species.      

The sphaeridia of sea urchins: ultrastructure and supposed function (Echinodermata,Echinoida)

Summary Sphaeridia are minute skeletal appendages of the echinoid test which are considered to be sense organs, organs of equilibrium, according to their shape. The sphaeridium forms a functional unit with the tubercle to which it adheres. The tubercle is encircled by a basiepithelial nerve ring of the epidermis. A circle of regularly arranged myocytes stretches from the tubercle to the sphaeridium. The muscles are distant from each other. The myofibrillar processes enter the pore space of the sphaeridial skeleton to which they are anchored by tendons; tendons are absent in the tubercle region. The cell bodies of the myocytes lie opposite to the nerve ring outside the skeleton. In this region the muscle cells and the nerve ring are in contact with each other, their basal laminae fuse. Tensions of the various myocytes are dependent on the position of the top-heavy sphaeridium. The nerve ring contains neurones which are provided with a cilium which lies close to the contact region with the myocytes. This arrangement leads to the assumption that the nerve cells in question have a proprioceptor function. Unique filter cells are present in the pore spaces of the sphaeridium and the tubercle. They possibly detoxicate the extracellular fluid that surrounds the myocytes. Phagocytes loaded with spacious phagosomes are crowded in the adjacent pore spaces. They are possibly extruded via the epidermis. Filter cells and phagocytes have obviously to do with the metabolism within the sphaeridium-tubercle-system.     

Embryonic expression of engrailed in sea urchins

Neural patterning genes that are expressed along the anterior-posterior axis of deuterostomes are expressed late in larval development in echinoderms and are thought to function in establishing the highly-derived, adult body plan. We have used genomic resources to clone an engrailed gene (SpEn) from Strongylocentrotus purpuratus, and with this we have developed an antibody specific for SpEn. SpEn is expressed late in embryogenesis in the developing larval nervous system. At the prism stage, a small number of neuroblasts in the oral ectoderm on the edge of the larval mouth begin expressing SpEn. The cells are in bilaterally symmetric positions. The expression of SpEn precedes the expression of the neural markers, synaptotagmin and serotonin in the SpEn immunoreactive cells. The SpEn cells are located on the margin of the domain of cells expressing SpNK2.1, but they do not have nuclear SpNK2.1. Expression of engrailed in a pair of bilateral neural structures in early development appears to be a shared feature of bilaterians.     

The evolution of repetitive DNA sequences in sea urchins.

Molecular hybridization of nuclear DNAs has been employed to study the evolution of the repetitive DNA sequences in four species of sea urchin. The data show that relative to S. purpuratus there has been approximately 0.1% sequence divergence per million years in the repetitive DNA sequences of S. droebachiensis, S. franciscanus, and L. pictus. These results confirm that repetitive DNA sequences are strongly conserved during evolution. However, comparison of the extent of base pair mismatch in the repetitive DNA heteroduplexes formed at Cot 20 with those formed at Cot 200 during the hybridization of S. purpuratus and L. pictus DNAs reveals that highly repetitive sequences of sea urchins may diverge more rapidly than do the more moderately repetitive sequences.     

Evolutionary reorganizations of ontogenesis in sea urchins

The data published during recent 15-20 years on comparative, experimental and molecular embryology of unusually developing sea urchins have been reviewed. These animals are characterized by large lipidrich eggs, highly modified embryogenesis, and the absence of a planktotrophic larva. Such a type of development is evolutionary advanced and arose independently in various phylogenetic lineages of the sea urchins.     

Heterochrony and heterotopy in the phylogeny of sea urchins

The role of heterochrony is evident in ontogeny and phylogeny of irregular (exocyclic) sea urchins. After metamorphosis, a juvenile passes the stage of regular (endocyclic) sea urchin, in which the periproct is surrounded by plates of the apical system. A shift of the periproct in the area of the fifth interambulacrum occurs in extant taxa at early stages of postlarval development and is accompanied by the reduction of genital plate 5. In some ancient (Jurassic) adult irregular sea urchins, the endocyclic state of the apical system is retained for a long time and the derivative of the fifth genital plate is sometimes observed even in Early Cretaceous species. Considerable transformations in the structure of the lower test surface in members of the order Spatangoida are manifested in changes in the relative positions of plastron plates and ambulacral areas I and V, separation of sternal plates from the labrum, etc. The mechanism of these changes is connected with translocation or “sliding” of sutures of particular plates as a result of nonuniform growth and partial resorption. The study of evolutionary lineages of Cretaceous and Cenozoic sea urchins has shown that the evolution was connected with the directional changes in some morphological characters at late ontogenetic stages. The process was accompanied by either extension, peramorphosis (lineages of the genera Micraster, Infulaster–Hagenowia in the European Province), or the loss of these stages, paedomorphosis (Hemiaster (Bolbaster) lineage, Late Eocene–Middle Miocene of Australia). The phenomena of heterochrony and heterotopy in the development of peripetal, marginal, and lateroanal fascioles in the Late Cretaceous and Paleocene families Hemiasteridae, Schizasteridae, and Paleopneustidae are described. The heterotopy is also illustrated by the example of the development of additional genital pores on ocular plates II and IV of the Middle Jurassic species Pygomalus analis (Disasteroida); its apical system has five pores instead of four. In the Late Cretaceous species Guettaria roccardi (Holasteroida), ocular plates II and IV have two pores each; in the apical system, there are eight genital pores instead of four. In some members of the order Holectypoida, the place of lost genital plate 5 is occupied by a new plate sometimes pierced by a pore, but judging from crystallographic data, it is not homologous to other genital plates. The order Clypeasteroida is characterized by the development of very small pores in both ambulacral and interambulacral fields; they provide passage for numerous accessory tube feet.     

The coelomic elements of sea urchins (Strongylocentrotus)

Summary The four coelomocyte classes of the red sea urchin,Strongylocentrotus franciscanus, described by light-microscope studies, are confirmed and the fine structure described. Material examined included fresh, non-aggregated cells; partially aggregated ones that had been heldin vitro up to four days; and aggregated cells heldin vitro for 40 days. Leukocytes from youngin-vitro preparations differed from most fresh leukocytes by having enlarged dense nucleoli and enlarged rough endoplasmic reticulum, which was often filled with secretion, and sometimes connected to the perinuclear cisterna. Leukocytes held 40 daysin vitro were mainly plasmodial. Unlike cells held a limited timein vitro, the 40-day leukocytes had nuclei much like those in fresh preparations.The three classes of spherule-bearing cells (vibratile cells, red spherule cells, and colorless spherule cells) differed greatly in ultrastructure, and varied in appearance according to the fixative and pH present during fixation. Vibratile-cell spherules were of biphasic construction, suggesting the condition of certain vertebrate mast cells. Red spherule cells occurred in two forms. The most common form in fresh preparations had despherulated,i.e., lacked material in the spherules; and the spherules of the second type were filled with either granular or homogeneous material. Colorless spherule cells had evenly and finely granular material in the spherules. Colorless spherule cells were uncommon or missing in material that had been heldin vitro. Certain unidentifiable spherule cells occurred in some preparations.Although samples are small, it is notable that in May and June, recognizable glycogen was present only in leukocytes that had been heldin vitro, not in any fresh cells. Glycogen occurred in fresh cells of all classes from samples taken in December and February (during or shortly before the normal spawning season ofS. franciscanus). Unlike the cells in fresh preparations made in May, June, and December, fresh leukocytes and vibratile cells taken in February often had extremely lobed nuclei and considerably developed rough endoplasmic reticulum.This investigation was supported by Public Health Service Research Grant No. 9296 (to P. T.Johnson) from the National Institute of Allergy and Infectious Diseases.     

The coelomic elements of sea urchins (Strongylocentrotus)

Summary Coelomocytes of Strongylocentrotus purpuratus and Strongylocentrotus franciscanus include phagocytic leukocytes, flagellated vibratile cells, and two types of spherule-bearing cells; one of which contains echinochrome, a red naphthaquinone pigment. Vibratile cells are PAS positive and contain a strongly acidic mucopolysaccharide. Substances with similar staining and chemical properties appear in the coelomic fluid under certain stress conditions. It is supposed that these substances enter the fluid by diffusion from the vibratile cells. Tests indicate that both spherule-cell types, like the spherule cells of holothurians (sea cucumbers), may undergo a cycle during which they pass from a basophilic to an acidophilic state. Chemical properties of the spherule cells approximate those of vertebrate mast cells. The variability of staining and chemical reactions of spherule cells explains why published accounts of their properties have differed so widely. It is suggested that the properties of mucoid substances in invertebrate spherule-bearing cells may be as variable as is true of vertebrate mast cells.This investigation was supported by Public Health Service Research Grant No. 5358 (to E. A. Steinhaus) from the National Institute of Allergy and Infectious Diseases.     

On the ultrastructure and the supposed function of the mineralizing matrix coat of sea urchins (Echinodermata,Echinoida)

Summary Echinoderm ossicles are part of the mesenchyme. Their formation and growth, with respect to the underlying tissues, is studied using echinoid spines and teeth and applying different methods of fixation. The calcification process in echinoderms is strictly intracellular and needs (1) syncytial sclerocytes which completely enclose (2) a vacuolar cavity which in turn contains (3) an organic matrix coat. Strictly speaking, each ossicle is nothing but the calcified vacuolar space of a single syncytium of sclerocytes. In fully grown parts, however, the continuous sheath may split open and the matrix-coated mineral may come into contact with the extracellular space. According to biochemical analyses the matrix consists of insoluble components, but most (95%) of its constituents are soluble in EDTA or weak acids. If routine transmission electron microscope methods are used the soluble components are lost and the matrix at best looks electron light. If tannic acid is added to the fixative the soluble matrix components are preserved and reveal further ultrastructural details of the biomineralization process in echinoderms. The matrix coat looks extremely electron dense. Further soluble material is to be found within the vacuolar space or attached to the vacuolar surface of the cytoplasmic sheath. The results lead to the opinion that the matrix coat consists of a hydrophobic framework of insoluble components that contains soluble components which guide the Ca through pores in the hydrophobic layers into the interior of the matrix-coated space. It is only within this space that the mineral is deposited.     

Nuclear activity during oogenesis in sea urchins

Summary Early meiotic stages of Arbacia punctulata oocytes have revealed the presence of synaptinemal complexes in the chromosomes, which persist through zygotene-pachytene. The synaptinemal complexes conform broadly to the usual tripartite structures found in other higher forms. In addition, nuclei at these stages consist of a small nucleolus and dense bodies of varying sizes. The nucleolus is fibrillar in texture throughout and does not seem to incorporate Uridine-5-³H after pulse labeling, whereas the chromosomes are labeled. The nucleolar label is visualized at diplotene stages and onwards. The nuclear envelope differentiates by the appearance of numerous nuclear pore complexes with dense material in the annuli, and the chromosomes become markedly diffused. At vitellogenesis stage the nucleolus and chromatin become highly labeled after pulse incorporation of Uridine, indicating synthesis of ribosomal and chromosomal RNAs.This investigation was supported by grants No. A-5049, A-3624 and D-17 from National Research Council, Canada, grant No. DRB-9340-05 (U6) from Defense Research Board, Canada, and grant No. DRG-918 AT from Damon Runyon Memorial Fund for Cancer Research.     

Glycerinated catch apparatus of sea urchin spines: the effects of cations on its mechanical properties and ultrastructure

Sea urchin spinal ligaments (the catch apparatus) were extracted with glycerin, and electron microscopic observations comfirmed that no cell membranes remained intact after glycerination. We studied the effects of cations (Na(+), K(+), Ca(2+), Mg(2+)) on the mechanical properties of the glycerinated ligaments. Monovalent cations decreased whereas divalent cations increased the viscosity of the ligaments. The ion dependencies were similar to previous results with detergent-extracted holothurian dermis, which suggests that the echinoid ligament shares a similar mechanism for changes in mechanical properties with other catch connective tissues. This provides evidence against the hypothesis of that muscles in the catch apparatus are responsible for the changes in mechanical properties of the ligament. Fine projections cross-bridging collagen fibrils were observed in the glycerin-extracted ligaments as well as in the intact ligaments. They were found in all the ionic conditions studied.     

Variability of sperm specific histones in sea urchins

The variability of sperm histones was compared in two species of sea urchin. Whole sperm specific histones (SpH), were isolated from Tetrapygus niger (Arbacoida) and Parechinus angulosus (Echinoida). Individual histones were purified by chromatography on BioGel P-60 followed by reverse high pressure liquid chromatography (HPLC). The heterogeneity of each major histone type from T. niger was established from their HPLC elution patterns and further confirmed by electrophoresis in polyacrylamide gels containing 6 mM Triton X-100 combined with a transverse urea gradient (0--8 M). In T. niger, as well as in P. angulosus, a single form of SpH1 and SpH2A were found. In contrast, SpH2B was found to be heterogeneous, but represented by one major form in both species. The relatedness between both sets of histones was determined by establishing their immunological cross-reactivity. In this context, polyclonal antibodies elicited against T. niger sperm histones were assayed against individual histones from P. angulosus. From the results obtained, it emerged that histone SpH2A was the more closely related protein between these two species, followed by histone SpH1. In contrast, histone SpH2B was found to be only moderately related. These results confirm that SpH2A did not co-evolve with SpH2B, as was predicted for most species.