Heliocidaris Tuberculata Research Articles

The role of visual and chemical cues in host detection by the symbiotic shrimp Gnathophylloides mineri

The use of visual and chemical cues in host detection is regarded as important but relatively unknown for many symbionts. The small circumtropical caridean shrimp Gnathophylloides mineri forms a close symbiotic association with sea urchins, particularly Tripneustes gratilla, in many parts of the world. G. mineri is known to occur in temperate eastern Australia but the breadth of host use and selection of hosts amongst different species of sea urchins is relatively unknown. The abundance of G. mineri on three co-occurring species of sea urchins, T. gratilla, Heliocidaris tuberculata and Pseudoboletia indiana were measured in eastern Australia. These species of sea urchins were chosen because of either a known prior association with this shrimp elsewhere in the world, or due to their abundance in the area in temperate Australia where the shrimp occurs. Field collections showed that the association between shrimp and sea urchin appears extremely host-specific, with G. mineri only observed on T. gratilla, at an average density of two shrimp per sea urchin. Moreover, the majority of symbionts occurred on the underside (oral region) of T. gratilla. A visual laboratory experiment showed that G. mineri would actively move towards T. gratilla in preference to the other potential host species in the absence of chemical signals. Using a Y-maze, we tested the reaction of G. mineri to the absence of visual signals but the presence of chemical signals originating from their host, and from other species of potential host sea urchins. Seawater alone was used as a negative control. We demonstrated that G. mineri could detect T. gratilla from a distance, the same host that the shrimp are associated with in the field, using visual and chemical cues. Moreover, G. mineri react quickly to visual and chemical host signals by moving towards T. gratilla in preference to other species of sea urchins or in the absence of a signal. The relative importance of visual versus chemical cues is, however, unknown.

Dissolved histamine: a potential habitat marker promoting settlement and metamorphosis in sea urchin larvae

Many species of marine invertebrate larvae settle and metamorphose in response to chemicals produced by organisms associated with the adult habitat, and histamine is a cue for larvae of the sea urchin Holopneustes purpurascens. This study investigated the effect of histamine on larval metamorphosis of six sea urchin species. Histamine induced metamorphosis in larvae of three lecithotrophic species (H. purpurascens, Holopneustes inflatus and Heliocidaris erythrogramma) and in one planktotrophic species (Centrostephanus rodgersii). Direct comparisons of metamorphic rates of lecithotrophic and planktotrophic larvae in assays cannot be made due to different proportions of larvae being competent. Histamine (10 μM) induced metamorphosis in 95% of larvae of H. purpurascens and H. inflatus after 1 h, while the coralline alga Amphiroa anceps induced metamorphosis in 40–50% of these larvae. Histamine (10 μM) and A. anceps induced 40 and 80% metamorphosis, respectively, in the larvae of H. erythrogramma after 24 h. Histamine (10 μM) and the coralline alga Corallina sp. induced 30 and 70% metamorphosis, respectively, in the larvae of C. rodgersii after 24 h. No metamorphosis of any larval species occurred in seawater controls. Larvae of two planktotrophic species (Tripneustes gratilla and Heliocidaris tuberculata) did not metamorphose in response to histamine. Seagrasses, the host plants of H. inflatus, induced rapid metamorphosis in larvae of the two Holopneustes species, and several algae induced metamorphosis in C. rodgersii larvae. Histamine leaching from algae and seagrasses may act as a habitat marker and metamorphic cue for larvae of several ecologically important sea urchin species.

Molecular phylogeny of echinometrid sea urchins: more species of Heliocidaris with derived modes of reproduction

Abstract. Pachechinus bajulus is an endemic Australian sea urchin with an unusual mode of brooded larval development. We used mitochondrial and nuclear gene sequences to estimate the phylogenetic relationships among Pachechinus and other Echinometridae, including well-studied species of Heliocidaris with planktonic development. We found strong evidence for the planktotrophic species Heliocidaris tuberculata as the sister group to a clade of three closely related species in which development is known (Heliocidaris erythrogramma, P. bajulus) or suspected (Pachechinus australiae) to be lecithotrophic. Clade support values and likelihood ratio tests rejected monophyly of Heliocidaris species. The sister group to H. erythrogramma is most likely the two Pachechinus species. We resolve the paraphyly problem by reassigning the Pachechinus species to the genus Heliocidaris (the senior synonym), which has six extant species including Heliocidaris australiae and Heliocidaris bajulus. The phylogeny has potentially important implications for comparative studies of developmental morphology and genetics that have assumed a close sister-group relationship between H. erythrogramma and H. tuberculata, and highlights the need for such data from H. bajulus and other Heliocidaris species.

Predicting water toxicity: Pairing passive sampling with bioassays on the Great Barrier Reef

Many coral reefs worldwide occur adjacent to urban or agricultural land which places these ecosystems at threat of exposure to complex mixtures of pollutants. In this study, the pairing of passive sampler extracts with bioassays is proposed as a tool for predicting effects of organic pollutant mixtures on key biota within coral reef ecosystems. Passive samplers, SDB-RPS Empore™ disks, which sequester a mixture of the contaminants present in the environment, were deployed at three sites in the Great Barrier Reef (GBR). Extracts from these samplers were analysed for herbicides and applied to bioassays targeting integral life stages or functions of coral reef biota. Biota included scleractinian coral larvae, sea urchin larvae, a marine diatom and marine bacteria. Photosynthesis in the marine diatom Phaeodactylum tricornutum was inhibited at the sampled environmental concentration while an environmental concentration factor of 15 times inhibited luminescence in the marine bacterium Vibrio fischeri. Concentrations of 50 times sampled environmental levels of organic pollutants inhibited >90% of Acropora millepora settlement and 100-fold environmental enrichment inhibited 100% Heliocidaris tuberculata larval development. These results demonstrate the utility of pairing passive sampling with bioassays and reveal that mixtures of organic pollutants in the GBR have the potential to cause detrimental effects to coral reef biota.

Overview of phenotypic plasticity in echinoid larvae, ‘Echinopluteus transversus’ type vs. typical echinoplutei

Many sea urchin echinoplutei exhibit phenotypic plasticity, increasing arm length to enhance food capture in nutrient poor conditions. We tested this phenomenon in species with contrasting larval forms reared in similar feeding conditions. Heliocidaris tuberculata has a typical echinopluteus larva with 4 pairs of arms while Centrostephanus rodgersii larvae have only 1 pair of arms (‘Echinopluteus transversus ’ type larva). Larvae were exposed to high, medium and no food treatments. The ratio ‘postoral arm length’ to ‘midline body length’ (PO:MBL) was used to document phenotypic plasticity. Fed H. tuberculata larvae developed short postoral arms and low PO:MBL, while starved larvae developed long postoral arms and high PO:MBL, indicative of plastic arm growth. In contrast, well fed C. rodgersii larvae had the longest arms and high PO:MBL, indicating the absence of plastic arm growth. Taking MBL into account, ANCOVA revealed that differences among treatments were due to PO. The hypothesis that starved C. rodgersii larvae would develop long arms was not supported. Principal component analysis confirmed that larvae in different food treatments had distinct morphologies and that H. tuberculata and C. rodgersii had opposite growth patterns with respect to food treatment. We suggest that ‘Echinopluteus transversus ’ type larvae are adapted for long distance dispersal, and with only 2 arms for feeding and swimming, there may be an imperative to maintain arm length irrespective of food conditions. Phenotypic plasticity in echinopluteal arm growth is not universal and may be influenced by phylogeny, latitude and the hydromechanics of larval form.

Direct toxicity assessment of volatile chlorinated hydrocarbon‐contaminated groundwater and derivation of a site‐specific guideline

Groundwater contaminated with a mixture of 14 volatile chlorinated hydrocarbons (VCHs) discharges to an estuarine embayment in Sydney, Australia. A screening-level hazard assessment identified a potential risk to aquatic organisms from surface water contaminated by the groundwater. Direct toxicity assessment of the groundwater was undertaken on 5 indigenous marine species to assess toxicity and derive a site-specific guideline. The testing included acute tests, subchronic tests on early life stages, and a chronic test. Test organisms included a microalga (Nitzschia closterium), an amphipod (Allorchestes compressa), a polychaete worm (Diopatra dentata), and sea urchin (Heliocidaris tuberculata) and oyster larvae (Saccostrea commercialis). Toxicity testing was undertaken in sealed containers to prevent loss of VCHs, and concentrations of VCHs were measured to accurately assess exposure concentrations. No observed effect concentration (NOEC) values varied from 1.56% dilution (1.11 mg total VCHs) to 50% dilution (45.5 mg total VCHs). EC50 values varied from 4.8% dilution (3.77 mg total VCHs) to more than 50% dilution (45.5 mg total VCHs). NOEC data were used to derive species sensitivity distributions (SSDs) and a site-specific guideline. SSDs were derived from Burr type III (including the Pareto) and log-normal distributions. The log-normal distribution represented the best fit, and because the Pareto distribution is a finite threshold model more suited to toxicants with a threshold mode of action, the log-normal SSD and the associated 95% trigger value (TV) of 830 microg/L of total VCHs, was adopted as the site-specific TV for the groundwater.

Toxicity of landfill leachate to sea urchin development with a focus on ammonia

Sea urchin gametes and embryos serve as a model system to evaluate toxicity in the marine environment. In this study, the toxicity of complex chemical mixtures in leachate samples to sea urchin development was examined with a focus on ammonia, which was the main contaminant of concern in most samples. Two rapid tests, the submitochondrial particle function and bacterial luminescence tests, were also used. Ammonia is highly toxic to sea urchin embryos with an EC50 of 1.3 mg l(-1) for the embryos of the Australian sea urchin Heliocidaris tuberculata. Leachate ammonia levels were well above these EC50 concentrations. To assess the contribution of ammonia to leachate toxicity in sea urchin development, we compared the predicted toxic units (PTU) and observed toxic units (OTU) for ammonia for each sample. The PTU/OTU comparison revealed that the sensitivity of the sea urchin embryos to ammonia were altered (enhanced or decreased) by other chemicals in the leachates. This result emphasises the need for parallel chemical analyses and a suite bioassays for evaluating the toxicity of complex and variable chemical mixtures.

Nodal expression and heterochrony in the evolution of dorsal–ventral and left–right axes formation in the direct‐developing sea urchin Heliocidaris erythrogramma

To understand the role of body axes in the evolution of larval form, we use the two sea urchins in the genus Heliocidaris, which have distinctly different larval morphologies. Heliocidaris tuberculata is an indirect-developing sea urchin, which forms a pluteus larva, whereas its sister species, Heliocidaris erythrogramma, exhibits direct development and forms a nonfeeding, ovoid larva. Changes along all three larval axes underlie the differences in larval form associated with each developmental mode. Nodal signaling has recently been implicated as important in establishing the dorsal-ventral (D-V) and left-right (L-R) axes in the indirect-developing sea urchin Paracentrotus lividus. However, because of changes in morphology and timing of morphogenetic events associated with the D-V and L-R axes, respectively, in H. erythrogramma, it was unclear whether nodal played the same roles during direct development. We show that the expression patterns and functions of nodal during H. erythrogramma development are similar to its roles in indirect-developing sea urchins in both D-V and L-R axes formation. However, there are profound changes in gene expression downstream of nodal signaling along the D-V axis and major heterochronies in the execution of the function of nodal along the L-R axis. These highly modified events are linked to the dramatic modifications of larval morphology that have occurred during the evolution of direct development in H. erythrogramma.

Co‐option and dissociation in larval origins and evolution: the sea urchin larval gut

The origin of marine invertebrate larvae has been an area of controversy in developmental evolution for over a century. Here, we address the question of whether a pelagic "larval" or benthic "adult" morphology originated first in metazoan lineages by testing the hypothesis that particular gene co-option patterns will be associated with the origin of feeding, indirect developing larval forms. Empirical evidence bearing on this hypothesis is derivable from gene expression studies of the sea urchin larval gut of two closely related but differently developing congenerics, Heliocidaris tuberculata (feeding indirect-developing larva) and H. erythrogramma (nonfeeding direct developer), given two subsidiary hypotheses. (1) If larval gut gene expression in H. tuberculata was co-opted from an ancestral adult expression pattern, then the gut expression pattern will remain in adult H. erythrogramma despite its direct development. (2) Genes expressed in the larval gut of H. tuberculata will not have a coordinated expression pattern in H. erythrogramma larvae due to loss of a functional gut. Five structural genes expressed in the invaginating archenteron of H. tuberculata during gastrulation exhibit substantially different expression patterns in H. erythrogramma with only one remaining endoderm specific. Expression of these genes in the adult of H. erythrogramma and larval gut of H. tuberculata, but not in H. erythrogramma larval endoderm, supports the hypothesis that they first played roles in the formation of adult structures and were subsequently recruited into larval ontogeny during the origin and evolution of feeding planktotrophic deuterostome larvae.

Gene expression patterns in a novel animal appendage: the sea urchin pluteus arm

The larval arms of echinoid plutei are used for locomotion and feeding. They are composed of internal calcite skeletal rods covered by an ectoderm layer bearing a ciliary band. Skeletogenesis includes an autonomous molecular differentiation program in primary mesenchyme cells (PMCs), initiated when PMCs leave the vegetal plate for the blastocoel, and a patterning of the differentiated skeletal units that requires molecular cues from the overlaying ectoderm. The arms represent a larval feature that arose in the echinoid lineage during the Paleozoic and offers a subject for the study of gene co-option in the evolution of novel larval features. We isolated new molecular markers in two closely related but differently developing species, Heliocidaris tuberculata and Heliocidaris erythrogramma. We report the expression of a larval arm-associated ectoderm gene tetraspanin, as well as two new PMC markers, advillin and carbonic anhydrase. Tetraspanin localizes to the animal half of blastula stage H. tuberculata and then undergoes a restriction into the putative oral ectoderm and future location of the postoral arms, where it continues to be expressed at the leading edge of both the postoral and anterolateral arms. In H. erythrogramma, its expression initiates in the animal half of blastulae and expands over the entire ectoderm from gastrulation onward. Advillin and carbonic anhydrase are upregulated in the PMCs postgastrulation and localized to the leading edge of the growing larval arms of H. tuberculata but do not exhibit coordinated expression in H. erythrogramma larvae. The tight spatiotemporal regulation of these genes in H. tuberculata along with other ontogenetic and phylogenetic evidence suggest that pluteus arms are novel larval organs, distinguishable from the processes of skeletogenesis per se. The dissociation of expression control in H. erythrogramma suggest that coordinate gene expression in H. tuberculata evolved as part of the evolution of pluteus arms, and is not required for larval or adult development.

Relationships Between Toxicity and Concentrations of Chemical Contaminants in Sediments from Sydney Harbour, Australia, and Vicinity

Correlation analyses between measures of toxicity and concentrations of chemical contaminants were conducted for 103 surficial sediments from Sydney Harbour, Australia, and vicinity. Toxicity tests consisted of amphipod survival and reburial tests of whole sediments (Corophium colo), sea urchin fertilisation and larval development tests of pore waters (Heliocidaris tuberculata) and microbial bioluminescence (Microtox) tests of solvent extracts and pore waters. Toxicity in most tests correlated with concentrations of metallic contaminants, in particular, zinc, lead and copper. Organic contaminants did not correlate as significantly with toxicity. However, Heliocidaris tuberculata showed relationships with organochlorine compounds in samples with low to moderate metals contamination. Toxicity in the Microtox solvent extract test appeared to be primarily influenced by the presence of sulfur. This study has no precedent in Australia and the results support the validity of using local indigenous species in toxicity tests of field-collected sediments. This toxicity/chemistry dataset may be used in evaluations of sediment quality guidelines recently introduced to Australia.

Dissociation of expression patterns of homeodomain transcription factors in the evolution of developmental mode in the sea urchins Heliocidaris tuberculata and H. erythrogramma

The direct-developing sea urchin species Heliocidaris erythrogramma has a radically modified ontogeny. Along with gains of novel features, its entire ectoderm has been reorganized, resulting in the apparent absence of a differentiated oral ectoderm, a major module present in the pluteus of indirect-developing species, such as H. tuberculata. The restoration of an obvious oral ectoderm in H. erythrogrammaxH. tuberculata hybrids, indicates the action of dominant regulatory factors from the H. tuberculata genome. We sought candidate regulatory genes based on the prediction that they should include genes that govern development of the oral ectoderm in the pluteus, but play different roles in H. erythrogramma. Such genes may have a large effect in the evolution of development. Goosecoid (Gsc), Msx, and the sea urchin Abd-B-like gene (Hox11/13b) are present and expressed in both species and the hybrid embryos. Both Gsc and Msx are oral ectoderm specific in H. tuberculata, and show novel and distinct expression patterns in H. erythrogramma. Gsc assumes a novel ectodermal pattern and Msx shifts to a novel and largely mesodermal pattern. Both Gsc and Msx show a restoration of oral ectoderm expression in hybrids. Hox11/13b is not expressed in oral ectoderm in H. tuberculata, but is conserved in posterior spatial expression among H. tuberculata, H. erythrogramma and hybrids, serving as a control. Competitive RT-PCR shows that Gsc, Msx, and Hox11/13b are under different quantitative and temporal controls in the Heliocidaris species and the hybrids. The implications for the involvement of these genes in the rapid evolution of a direct developing larva are discussed.

Major regulatory factors in the evolution of development: the roles of goosecoid and Msx in the evolution of the direct‐developing sea urchin Heliocidaris erythrogramma

The transcription factors Gsc and Msx are expressed in the oral ectoderm of the indirect-developing sea urchin Heliocidaris tuberculata. Their patterns of expression are highly modified in the direct developer Heliocidaris erythrogramma, which lacks an oral ectoderm. We here test the hypothesis that they are large effect genes responsible for the loss of the oral ectoderm module in the direct-developing larva of H. erythrogramma as well as for the restoration of an overt oral ectoderm in H.e. xH.t. hybrids. We undertook misexpression/overexpression and knockdown assays in the two species and in hybrids by mRNA injection. The results indicate that dramatic changes of function of these transcription factors has occurred. One of these genes, Gsc, has the ability when misexpressed to partially restore oral ectoderm in H. erythrogramma. On the other hand, Msx has lost any oral function and instead has a role in mesoderm proliferation and patterning. In addition, we found that the H. tuberculataGsc is up regulated in H.e. xH.t. hybrids, showing a preferential use of the indirect developing parental gene in the development of the hybrid. We suggest that Gsc qualifies as a gene of large evolutionary effect and is partially responsible for the evolution of direct development of H. erythrogramma. We discuss these results in light of modularity and genetic networks in development, as well as in their implications for the rapid evolution of large morphological changes in development.

Larval ectoderm, organizational homology, and the origins of evolutionary novelty

Comprehending the origin of marine invertebrate larvae remains a key domain of research for evolutionary biologists, including the repeated origin of direct developmental modes in echinoids. In order to address the latter question, we surveyed existing evidence on relationships of homology between the ectoderm territories of two closely related sea urchin species in the genus Heliocidaris that differ in their developmental mode. Additionally, we explored a recently articulated idea about homology called 'organizational homology' (Müller 2003. In: Müller GB, Newman SA, editors. Origination of organismal form: beyond the gene in developmental and evolutionary biology. Cambridge, MA: A Bradford Book, The MIT Press. p 51-69. ) in the context of this specific empirical case study. Applying the perspective of organizational homology to our experimental system of congeneric echinoids has led us to a new hypothesis concerning the ectoderm evolution in these species. The extravestibular ectoderm of the direct developer Heliocidaris erythrogramma is a novel developmental territory that arose as a fusion of the oral and aboral ectoderm territories found in indirect developing echinoids such as Heliocidaris tuberculata. This hypothesis instantiates a theoretical principle concerning the origin of developmental modules, 'integration', which has been neglected because the opposite theoretical principle, 'parcellation', is more readily observable in events such as gene duplication and divergence (Wagner 1996. Am Zool 36:36-43).

Morphological evolution in sea urchin development: hybrids provide insights into the pace of evolution.

Hybridisations between related species with divergent ontogenies can provide insights into the bases for evolutionary change in development. One example of such hybridisations involves sea urchin species that exhibit either standard larval (pluteal) stages or those that develop directly from embryo to adult without an intervening feeding larval stage. In such crosses, pluteal features were found to be restored in fertilisations of the eggs of some direct developing sea urchins (Heliocidaris erythrogramma) with the sperm of closely (Heliocidaris tuberculata) and distantly (Pseudoboletia maculata) related species with feeding larvae. Such results can be argued to support the punctuated equilibrium model-conservation in pluteal regulatory systems and a comparatively rapid switch to direct development in evolution.1,2 Generation of hybrids between distantly related direct developers may, however, indicate evolutionary convergence. The 'rescue' of pluteal features by paternal genomes may require maternal factors from H. erythrogramma because the larva of this species has pluteal features. In contrast, pluteal features were not restored in hybridisations with the eggs of Holopneustes purpurescens, which lacks pluteal features. How much of pluteal development can be lost before it cannot be rescued in such crosses? The answer awaits hybridisations among indirect and direct developing sea urchins differing in developmental phenotype, in parallel with investigations of the genetic programs involved.

Evolution of OTP‐Independent larval skeleton patterning in the direct‐developing sea urchin, Heliocidaris erythrogramma

Heliocidaris erythrogramma is a direct-developing sea urchin that has evolved a modified ontogeny, a reduced larval skeleton, and accelerated development of the adult skeleton. The Orthopedia gene (Otp) encodes a homeodomain transcription factor crucial in patterning the larval skeleton of indirect-developing sea urchins. We compare the role of Otp in larvae of the indirect-developing sea urchin Heliocidaris tuberculata and its direct-developing congener H. erythrogramma. Otp is a single-copy gene with an identical protein sequence in these species. Expression of Otp is initiated by the late gastrula, initially in two cells of the oral ectoderm in H. tuberculata. These cells are restricted to oral ectoderm and exhibit left-right symmetry. There are about 266 copies of Otp mRNA per Otp- expressing cell in H. tuberculata. We tested OTP function in H. tuberculata and H. erythrogramma embryos by microinjection of Otp mRNA. Mis-expression of Otp mRNA in H. tuberculata radialized the embryos and caused defects during larval skeletogenesis. Mis-expression of Otp mRNA in H. erythrogramma embryos did not affect skeleton formation. This is consistent with the observation by in situ hybridization of no concentration of Otp transcript in any particular cells or region of the H. erythrogramma larva, and measurement of a level of less than one copy of endogenous Otp mRNA per cell in H. erythrogramma. OTP plays an important role in patterning the larval skeleton of H. tuberculata, but this role apparently has been lost in the evolution of the H. erythrogramma larva, and replaced by a new patterning mechanism.

Assessment of metal toxicity in sediment pore water from Lake Macquarie, Australia.

Recent investigations into the level of heavy metal enrichment in the sediments of Lake Macquarie have indicated that significant contamination has occurred over the past 100 years, with elevated levels of lead, zinc, cadmium, copper, and selenium being observed in most parts of the lake. Pore water extracted from sediments showing the greatest contamination by these metals exhibited toxicity to the larval development of the sea urchin Heliocidaris tuberculata. However, an analysis of pore water metal concentrations revealed that the concentrations of these metals were too low to cause toxicity. Rather, pore water toxicity was highly correlated with manganese for the majority of sites sampled; subsequent spiking experiments confirmed manganese as a cause of toxicity. Current levels of manganese in the sediments of Lake Macquarie have arisen from natural sources and are not the result of anthropogenic activities. These results reiterate the importance of identifying the causes of toxicity in assessments of sediment contamination, particularly when testing sediment pore waters using sensitive early life stages.

ADAPTIVE EVOLUTION OF BINDIN IN THE GENUS HELIOCIDARIS IS CORRELATED WITH THE SHIFT TO DIRECT DEVELOPMENT

Sea urchins are widely used to study both fertilization and development. In this study we combine the two fields to examine the evolution of reproductive isolation in the genus Heliocidaris. Heliocidaris tuberculata develops indirectly via a feeding larva, whereas the only other species in the genus, H. erythrogramma, has evolved direct development through a nonfeeding larva. We estimated the time of divergence between H. erythrogramma and H. tuberculata from mitochondrial DNA divergence, quantified levels of gametic compatibility between the two species in cross-fertilization assays, and examined the mode of evolution of the sperm protein bindin by sequencing multiple alleles of the two species. Bindin is the major component of the sea urchin sperm acrosomal vesicle, and is involved in sperm-egg attachment and fusion. Based on our analyses, we conclude that: the two species of Heliocidaris diverged less than five million years ago, indicating that direct development can evolve rapidly in sea urchins; since their divergence, the two species have become gametically incompatible; Heliocidaris bindin has evolved under positive selection; and this positive selection is concentrated on the branch leading to H. erythrogramma. Three hypotheses can explain the observed pattern of selection on bindin: (1) it is a correlated response to the evolution of direct development in H. erythrogramma; (2) it is the result of an intraspecific process acting in H. erythrogramma but not in H. tuberculata; or (3) it is the product of reinforcement on the species that invests more energy into each egg to avoid hybridization.

Novel gene expression patterns in hybrid embryos between species with different modes of development.

Cross-species hybrids between eggs of the direct-developing sea urchin, Heliocidaris erythrogramma, and sperm from its congeneric indirect-developing species, Heliocidaris tuberculata, show restoration of features of the paternal feeding pluteus larva, including the gut, and pluteus spicular skeleton. Unlike other reported sea urchin cross-species hybrids, Heliocidaris hybrids express genes derived from both maternal and paternal species at high levels. Ectodermal cell types, which differ radically between the two parental species, are of intermediate form in the hybrids. Gene expression patterns in hybrid embryo tissues represent a number of combinations of parental gene expression patterns: genes that are not expressed in one paternal species, but are expressed in hybrids as in the expressing parent; genes that show additive expression patterns plus novel sites of expression; a gene that is misexpressed in the hybrids; and genes expressed identically in both parents and in hybrids. The results indicate that both conserved and novel gene regulatory interactions are present. Only one gene, CyIII actin, has lost cell-type-specific regulation in the hybrids. Hybrids thus reveal that disparate parental genomes, each with its own genic regulatory system, can produce in combination a novel gene expression entity with a unique ontogeny. This outcome may derive from conserved gene regulatory regions in downstream genes of both parental species responding in conserved ways to higher-level regulators that determine modular gene expression territories.

Modularity and dissociation in the evolution of gene expression territories in development.

Modularity is a salient feature of development and crucial to its evolution. This paper extends modularity to include the concept of gene expression territory, as established for sea urchin embryos. Territories provide a mechanism for partitioning of the cells of a rapidly developing embryo into functional units of a feeding larva. Territories exhibit the characteristics of modules. The paper asks if the embryo and the nonfeeding larva of the direct-developing sea urchin Heliocidaris erythrogramma are organized into gene expression territories, and if its territories correspond to the canonical territories of the pluteus. An analysis of cell lineage and gene expression data for H. erythrogramma shows that skeletogenic cell, coelomic, and vegetal plate gene expression territories are conserved, although they arise from cell lineages distinct from those of the pluteus, and the overall morphology of the larva differs from that of a pluteus. The ectoderm, as in indirect developers, is divided into territories. However, the oral ectodermal territory characteristic of the pluteus is absent in H. erythrogramma. Oral ectoderm is restored in hybrids of H. erythrogramma eggs fertilized by Heliocidaris tuberculata sperm. This indicates that embryonic modules evolve by changes in expression of dominant regulatory genes within territories and that entire modules can be eliminated in evolution of embryos.