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R A Raff

Publications and source records attributed to R A Raff.

At least 37 records · Page 2Linked to original sources
PubMed1995

A sea urchin homologue of ceh-19, an unusual homeobox-containing gene from a nematode.

When screening for homeobox-containing genes from the sea urchin Heliocidaris erythrogramma (He), we isolated an exon of a gene which appears to be a homologue of the homeobox-containing gene, ceh-19, of Caenorhabditis elegans (Ce). The predicted translation of the sea urchin sequence shows 77% identity and 92% similarity to the first 53 amino acids of the homeodomain of ceh-19. The ceh-19 gene exhibits an intron in an unusual location in the 3' end of the homeobox; the He gene shares this feature.

Amino Acid Sequence↗
PubMed1995

Structure and evolution of CyI cytoplasmic actin-encoding genes in the indirect- and direct-developing sea urchins Heliocidaris tuberculata and Heliocidaris erythrogramma.

The CyI cytoplasmic actin-encoding genes of Heliocidaris erythrogramma (He), a direct-developing sea urchin, and H. tuberculata, an indirect developer, were isolated and compared to the homologous CyI gene of another indirect developer, Strongylocentrotus purpuratus. Comparisons show that despite the differences in development, the actin gene structures and sequences are highly similar. The coding and 3' untranslated regions are conserved. The 5' He regulatory region has an inserted repeat element, but is otherwise similar to its homologues in the arrangement of presumptive transcription control elements.

Actins↗
PubMed1995

Structure, expression, and extracellular targeting of PM27, a skeletal protein associated specifically with growth of the sea urchin larval spicule.

The PM27 gene encodes a sea urchin skeletal protein. Both the transcript and encoded protein appear at the mesenchyme blastula stage and are restricted to the primary mesenchyme cell (PMC) lineage throughout development. Transgenic expression of PM27 promoter constructs demonstrates that this cell specificity is regulated at the level of transcription. The PM27 sequence predicts a nonglycosylated secretory product of 27 kDa in mature form. The N-terminal "repeat" domain of the deduced amino acid sequence consists of a series of tandem repeats and shares both sequence and predicted structural similarities with several fiber-forming proteins. The C-terminal "lectin" domain is similar to the C-class lectins. Antisera against either domain of PM27 detect two major proteins in embryo extracts, with apparent molecular weights of 27 and 30 kDa. Immunolocalization in whole embryos demonstrates that PM27 antigen is produced uniformly and exclusively by PMCs through the early prism stage and that this specificity is further restricted during skeletogenesis to a subpopulation of PMCs associated with the growing tips of the spicules. It is secreted to the skeletal compartment and accumulates predominantly at the advancing mineralizing surface of the spicule tips. As the spicules elongate PM27 protein disappears from the more mature mid-shaft regions. The observed characteristics of PM27 are consistent with a role in the regulation or execution of skeletal growth, as opposed to maintenance or structural integrity of the spicules.

Amino Acid Sequence↗
PubMed1995

Evolution of the fibropellin gene family and patterns of fibropellin gene expression in sea urchin phylogeny.

This study documents evolutionary modifications in the expression patterns of the sea urchin EGF I and EGF III genes, which encode a family of extracellular matrix proteins, the fibropellins. We show that the sea urchin apical lamina, a macromolecular extracellular matrix that surrounds the sea urchin embryo and is made up of the fibropellins, has been conserved through at least 250 million years of echinoid evolution. The contribution of different fibropellin family members to this structure has, however, changed over the course of sea urchin phylogeny, and between two congeneric species that exhibit different developmental modes. Mapping the evolutionary history of the EGF genes on a cladogram of relationships among sea urchins reveals that EGF I is present in all echinoids examined, while EGF III appears to have arisen by duplication and divergence from EGF I during the radiation of a suborder of the camarodont sea urchins some 35-45 million years ago. Alterations in the temporal expression patterns of these genes as well as the loss of one of the two EGF I transcripts and encoded protein are coincident with the evolution of a direct-developing larval form in Heliocidaris erythrogramma. H. erythrogramma and its congener Heliocidaris tuberculata, which develops via a typical echinopluteus larva, shared a common ancestor about 10 million years ago. The differences in fibropellin representation within the apical lamina of the various taxa indicate that a homologous embryonic structure can undergo substantial changes in composition during its evolutionary history.

Animals↗
PubMed1994

Dollo's law and the death and resurrection of genes.

Dollo's law, the concept that evolution is not substantively reversible, implies that the degradation of genetic information is sufficiently fast that genes or developmental pathways released from selective pressure will rapidly become nonfunctional. Using empirical data to assess the rate of loss of coding information in genes for proteins with varying degrees of tolerance to mutational change, we show that, in fact, there is a significant probability over evolutionary time scales of 0.5-6 million years for successful reactivation of silenced genes or "lost" developmental programs. Conversely, the reactivation of long (> 10 million years)-unexpressed genes and dormant developmental pathways is not possible unless function is maintained by other selective constraints; the classic example of the resurrection of "hen's teeth" is most likely an experimental artifact, and the experimental reactivation of the Archaeopteryx limb developmental program has been shown to be a misinterpretation. For groups undergoing adaptive radiations, lost features may "flicker" on and off, resulting in a distribution of character states that does not reflect the phylogeny of the group.

Ambystoma mexicanum↗
PubMed1994

Deuterostome phylogeny and the sister group of the chordates: evidence from molecules and morphology.

Complete coding regions of the 18S rRNA gene of an enteropneust hemichordate and an echinoid and ophiuroid echinoderm were obtained and aligned with 18S rRNA gene sequences of all major chordate clades and four outgroups. Gene sequences were analyzed to test morphological character phylogenies and to assess the strength of the signal. Maximum-parsimony analysis of the sequences fails to support a monophyletic Chordata; the urochordates form the sister taxon to the hemichordates, and together this clade plus the echinoderms forms the sister taxon to the cephalochordates plus craniates. Decay, bootstrap, and tree-length distribution analyses suggest that the signal for inference of dueterostome phylogeny is weak in this molecule. Parsimony analysis of morphological plus molecular characters supports both monophyly of echinoderms plus enteropneust hemichordates and a sister group relationship of this clade to chordates. Evolutionary parsimony does not support chordate monophyly. Neighbor-joining, Fitch-Margoliash, and maximum-likelihood analyses support a chordate lineage that is the sister group to an echinoderm-plus-hemichordate lineage. The results illustrate both the limitations of the 18S rRNA molecule alone for high-level phylogeny inference and the importance of considering both molecular and morphological data in phylogeny reconstruction.

Animals↗
PubMed1993

The SpEGF III gene encodes a member of the fibropellins: EGF repeat-containing proteins that form the apical lamina of the sea urchin embryo.

We have identified a gene in the sea urchin Strongylocentrotus purpuratus that encodes a protein with multiple epidermal growth factor(EGF)-like motifs. The SpEGF III cDNA sequence predicts a 570 amino acid protein with a complex domain structure similar to that of the fibropellins--the protein products of the SpEGF I gene. A putative hydrophobic leader sequence is followed by an EGF-like motif, a domain similar to complement Cls, seven tandem EGF-like repeats, and a carboxy terminal domain similar to avidin. The 2.9-kb SpEGF III mRNA is expressed from a single copy gene. SpEGF III mRNA is present at low levels in unfertilized eggs and during early cleavage, then rapidly increases in abundance between late morula and mesenchyme blastula stages to maximal levels maintained through subsequent stages. Polyclonal antisera from SpEGF III fusion proteins reveal that the protein is a 100-kDa fibropellin which is present in unfertilized eggs but does not accumulate substantially until the mesenchyme blastula stage. The protein is localized to the apical lamina, a structurally complex component of the extracellular matrix that is made up of three major proteins, two of which are the differentially spliced products of SpEGF I. The size and localization of the SpEGF III protein, and the results of immunoprecipitation assays which reveal that it is tightly associated with the products of SpEGF I, indicate that it is the third major protein component of the apical lamina. The timing of SpEGF III protein accumulation is coincident with an increase in the structural complexity of the apical lamina, and with the developmental period when the apical lamina plays an important role in gastrulation.

Amino Acid Sequence↗
PubMed1993

Protein-DNA interactions at putative regulatory regions of two coordinately expressed genes, msp130 and PM27, during skeletogenesis in sea urchin embryos.

Development of the primary mesenchyme cells (PMCs) of the sea urchin embryo, which give rise to the larval skeleton, involves the coordinate onset of expression of several structural genes. As part of an effort to identify cis-acting elements that might play a role in this regulatory event, co-regulated genes were examined by two approaches. First, they were compared for conserved sequence elements. Four conserved elements were found as a cluster in all three genes examined, suggesting a regulatory role. Second, as a test for potential function, the putative regulatory regions of two of these genes were examined for protein binding sites. DNase I protection and gel mobility shift assays were used to: 1) identify several nuclear protein binding sites in these regions, two of which correspond to conserved elements among the genes; 2) demonstrate that the developmental time of appearance of the proteins that interact with these sites corresponds to the time of activation of the genes; and 3) show that two of the conserved sequence elements shared by these genes compete for the same binding proteins. These data identify putative regulatory elements, whose specific roles in the coordinate regulation of PMC-expressed genes can now be addressed directly using appropriate transgenic expression constructs.

Animals↗
PubMed1992

Direct-developing sea urchins and the evolutionary reorganization of early development.

The evolution of development can be made accessible to study by exploiting closely related species that exhibit distinct ontogenies. The direct-developing sea urchin Heliocidaris erythrogramma is closely related to indirect-developing sea urchins that develop via a feeding larval stage. Superficial consideration would suggest that simple heterochronies resulting in loss of larval features and acceleration of adult features could explain the substitution of direct for indirect development. However, our experiments show that early development has in fact been extensively remodeled, with modified localization of maternal determinants coupled with dissociation of cell cleavage from axis formation resulting in novel patterns of cell lineage differentiation and fate map. Gene expression has undergone concomitant changes.

Animals↗
PubMed1992

Phylogenetic position of phylum Nemertini, inferred from 18S rRNA sequences: molecular data as a test of morphological character homology.

Partial 18S rRNA sequence of the nemertine Cerebratulus lacteus was obtained and compared with those of coelomate metazoans and acoelomate platyhelminths to test whether nemertines share a most recent common ancestor with the platyhelminths, as traditionally has been implied, or whether nemertines lie within a protostome coelomate clade, as suggested by more recent morphological analyses. Maximum-parsimony analysis supports the inclusion of the nemertine within a protostome-coelomate clade that falls within a more inclusive coelomate clade. Bootstrap analysis indicates strong support for a monophyletic Coelomata composed of a deuterostome and protostome-coelomate clade. Support for a monophyletic protostome Coelomata is weak. Inference by distance analysis is consistent with that of maximum parsimony. Analysis of down-weighted paired sites by maximum parsimony reveals variation in topology only within the protostome-coelomate clade. The relationships among the protostome coelomates cannot be reliably inferred from the partial sequences, suggesting that coelomate protostomes diversified rapidly. Results with evolutionary parsimony are consistent with the inclusion of the nemertine in a coelomate clade. The molecular inference corroborates recent morphological character analyses that reveal no synapomorphies of nemertines and flatworms but instead suggest that the circulatory system and rhynchocoel of nemertines are homologous to coelomic cavities of protostome coelomates, thus supporting the corresponding hypothesis that nemertines belong within a protostome-coelomate clade. The sequence data provide an independent test of morphological character homology.

Animals↗
PubMed1992

Evolutionary dissociation between cleavage, cell lineage and embryonic axes in sea urchin embryos.

Using vital dye staining and the microinjection of fluorescent cell lineage-autonomous tracers, the relationship between the first cleavage plane and the prospective larval dorsoventral axis was examined in several sea urchin species, including: Strongylocentrotus purpuratus, S. droebachiensis, Lytechinus pictus, Clypeaster rosaceus, Heliocidaris tuberculata and H. erythrogramma. The results indicate that there is no single relationship between the early cleavage pattern and the dorsoventral axis for all sea urchins; however, specific relationships exist for individual species. In S. purpuratus the first cleavage plane occurs at an angle 45 degrees clockwise with respect to the prospective dorsoventral axis in most cases, as viewed from the animal pole. On the other hand, in S. droebachiensis, L. pictus and H. tuberculata, the first cleavage plane generally corresponds with the plane of bilateral symmetry. There does not appear to be a predominant relationship between the first cleavage plane and the dorsoventral axis in C. rosaceus. In the direct-developing sea urchin H. erythrogramma the first cleavage plane bisects the dorsoventral axis through the frontal plane. Clearly, evolutionary differences have arisen in the relationship between cleavage pattern and developmental axes. Therefore, the mechanism of cell determination is not necessarily tied to any particular pattern of cell cleavage, but to an underlying framework of axial systems resident within sea urchin eggs and embryos.

Animals↗
PubMed1992

Evolution of developmental decisions and morphogenesis: the view from two camps.

Modern developmental biology largely ignores evolution and instead focuses on use of standard model organisms to reveal general mechanisms of development. Evolutionary biologists more widely hold developmental biology to be of major consequence in providing potential insights into evolution. Evolutionary insights can enlighten our views of developmental mechanisms as much as developmental data offer clearer views of mechanisms which underlie evolutionary change. However, insights have been limited by the long-term disengagement of the two fields dating to the rise of experimental embryology in the 1890s. Molecular genetics now provides a powerful tool to probe both gene function and evolutionary relationships, and a greater connection has become possible. The expansion of experimental organisms beyond the standard model animals used in most studies of development allows us to ask deeper questions about the interaction of development and evolution. This paper presents an analysis of the complementary uses of the resulting data in the two fields as they grope for accommodation. Analysis of the radical changes in early development seen in closely related sea urchins with alternate modes of development illustrate the complementarity of developmental and evolutionary data. These studies show that what have been thought to be constrained mechanisms of axial determination, cell lineage patterning, and gastrulation in fact evolve readily and provide the means for the rapid evolution of development.

Animals↗
PubMed1991

Fibropellins, products of an EGF repeat-containing gene, form a unique extracellular matrix structure that surrounds the sea urchin embryo.

The sea urchin SpEGF 1 gene belongs to a growing family of developmentally important genes which encode proteins that contain repeated epidermal growth factor-like motifs. To characterize the embryonic expression of the protein products of this gene from Strongylocentrotus purpuratus, we generated polyclonal antisera from SpEGF I fusion proteins. These antibodies recognize two glycoproteins of 145 and 185 kDa, which we have named fibropellins. These proteins are present in unfertilized oocytes and throughout early development. The fibropellins are stored in cytoplasmic vesicles in the oocyte and are released soon after fertilization in a distinct secretory event following the exocytosis of cortical granule contents. Following secretion the proteins are localized in the basal surface of the hyaline layer. At the blastula stage the fibropellins become organized into distinct fibers which form a mesh-like network over the surface of the embryo. During subsequent development to the pluteus larva stage this network increases in overall morphological complexity and becomes regionally distinct. The molecular weights of the fibropellins and their pattern of embryonic localization indicate that these proteins form a component of the hyaline layer previously described as the apical lamina.

Animals↗
PubMed1991

The phylogenetic status of arthropods, as inferred from 18S rRNA sequences.

Partial 18S rRNA sequences of five chelicerate arthropods plus a crustacean, myriapod, insect, chordate, echinoderm, annelid, and platyhelminth were compared. The sequence data were used to infer phylogeny by using a maximum-parsimony method, an evolutionary-distance method, and the evolutionary-parsimony method. The phylogenetic inferences generated by maximum-parsimony and distance methods support both monophyly of the Arthropoda and monophyly of the Chelicerata within the Arthropoda. These results are congruent with phylogenies based on rigorous cladistic analyses of morphological characters. Results support the inclusion of the Arthropoda within a spiralian or protostome coelomate clade that is the sister group of a deuterostome clade, refuting the hypothesis that the arthropods represent the "primitive" sister group of a protostome coelomate clade. Bootstrap analyses and consideration of all trees within 1% of the length of the most parsimonious tree suggest that relationships between the nonchelicerate arthropods and relationships within the chelicerate clade cannot be reliably inferred with the partial 18S rRNA sequence data. With the evolutionary-parsimony method, support for monophyly of the Arthropoda is found in the majority of the combinations analyzed if the coelomates are used as "outgroups." Monophyly of the Chelicerata is supported in most combinations assessed. Our analyses also indicate that the evolutionary-parsimony method, like distance and parsimony, may be biased by taxa with long branches. We suggest that a previous study's inference of the Arthropoda as paraphyletic may be the result of (a) having two few arthropod taxa available for analysis and (b) including long-branched taxa.

Animals↗
PubMed1990

Promoter structure and protein sequence of msp130, a lipid-anchored sea urchin glycoprotein.

The early fate specification of primary mesenchyme cells in sea urchin embryos makes them an attractive system for studying alterations in gene expression and protein synthesis during cell lineage determination and differentiation. To analyze the developmental regulation of gene expression in Strongylocentrotus purpuratus, we have isolated and sequenced genomic and cDNA clones encoding msp 130, a mesenchyme-specific cell surface glycoprotein. We have located the transcription initiation site of the msp130 gene and sequenced several kilobases of the promoter region. The region of the gene that encodes the protein is divided into numerous small (less than 500 base pairs) exons. The msp130 protein possesses two novel glycine-rich domains and a signal peptide, but apparently lacks a transmembrane domain. The carboxyl-terminal sequence suggests that msp130 may be phosphatidylinositol-linked to the cell membrane, and experiments with phospholipases support this conclusion. The implications of the msp130 sequence for its possible functions are discussed.

Amino Acid Sequence↗
PubMed1990

Novel origins of lineage founder cells in the direct-developing sea urchin Heliocidaris erythrogramma.

The lineage and fate of each blastomere in the 32-cell embryo of the direct-developing sea urchin Heliocidaris erythrogramma have been traced by microinjection of tetramethylrhodamine-dextran. The results reveal substantive evolutionary modifications of the ancestral cell lineage pattern of indirect sea urchin development. Significant among these modifications are changes in the time and order of cell lineage segregation: vegetal ectodermal founder cells consistently arise earlier than during indirect development, while internal founder cells generally segregate later and in a different sequence. Modifications have also arisen in proportions of the embryo fated to become various cell types and larval structures. Ectodermal fates, particularly vestibular ectoderm, comprise a greater proportion of the total cellular volume in H. erythrogramma. Among internal cell types, coelom consumes more and endoderm less of the remaining cellular volume than during indirect sea urchin development. Evolutionary modifications are also apparent in the positional origin of larval cell types and structures in H. erythrogramma. These include an apparent tilt in the axis of prospective cell fate relative to the animal-vegetal axis as defined by cleavage planes. Together these evolutionary changes in the cell lineage of H. erythrogramma produce an accelerated loss of dorsoventral symmetry in cell fate relative to indirect development. The extent and diversity of rearrangements in its cell lineage indicate that the non-feeding larva of H. erythrogramma is a highly modified, novel form rather than a degenerate pluteus larva. These same modifications underscore the evolutionarily flexible relationship between cell lineage, gene expression, and larval morphology in sea urchin development.

Animals↗
PubMed1990

Evolutionary change in the process of dorsoventral axis determination in the direct developing sea urchin, Heliocidaris erythrogramma.

Embryos of the indirect developing sea urchin, Heliocidaris tuberculata, and of Heliocidaris erythrogramma which develops directly without the formation of a pluteus larva, were bisected at the two- and four-cell stages. Paired half-embryos resulting from the bisection of H. tuberculata embryos along either the first or the second cleavage plane develop identically into miniature prism stage larvae. As in other indirect developing sea urchins, no differential segregation of developmental potential takes place as a result of the first and second cleavage divisions. Although half-embryos resulting from bisection along the second cleavage plane differentiate all cell types and develop equivalently in H. erythrogramma, the isolated first cleavage blastomeres do not. One of these two cells always forms significantly more mesodermal and endodermal cells. These patterns of differentiation are consistent with fate-mapping studies indicating that most mesodermal and endodermal cells are derived from the prospective ventral blastomere. Therefore, a differential segregation of developmental potential takes place at the first cleavage division in H. erythrogramma. When embryos of H. erythrogramma were bisected during the eight-cell stage, isolated tiers of animal blastomeres typically formed only ectodermal structures including the vestibule, whereas vegetal embryo halves formed all differentiated cell types. We propose that animal-vegetal cell determination and differentiation takes place along an axis which has been shifted relative to the pattern of cell cleavages in the embryos of H. erythrogramma. Vegetal morphogenetic potential for the formation of mesodermal and endodermal structures has become more closely associated with the prospective ventral side of the embryo during the evolution of direct development in Heliocidaris.

Animals↗