Molecular Embryology Research Articles

Insights from sharks: Evolutionary and developmental models of fin development

The shark and its embryology have recently returned to the spotlight as a model animal in the quest to determine the origins of paired appendages during vertebrate evolution. As the most basal living gnathostomes, sharks and other extant chondrichthyans are ideal models to elucidate the developmental mechanisms utilised in mesoderm-derived primitive fin morphologies. Chondrichthyans occupy a phylogenetic position and possess morphological structures that can answer major questions on the origin of the body plan of vertebrates. This review will outline the past, present, and future use of shark species as a model system with particular emphasis on the recent studies that have utilised comparative molecular embryology of chondrichthyan species to examine the question of the origin of the paired fins. We will also examine the problems and pitfalls of utilising chondrichthyans and the barriers that remain to their utilisation in the modern era of developmental biology.

Hagfish embryology with reference to the evolution of the neural crest

Hagfish, which lack both jaws and vertebrae, have long been the subject of intense interest owing to their position at a crucial point in the evolutionary transition to a truly vertebrate body plan. However, unlike the comparatively well characterized vertebrate agnathan lamprey, little is known about hagfish development. The inability to analyse hagfish at early embryonic stages has frustrated attempts to resolve questions with important phylogenetic implications, including fundamental ones relating to the emergence of the neural crest. Here we report the obtainment of multiple pharyngula-stage embryos of the hagfish species Eptatretus burgeri and our preliminary analyses of their early development. We present histological evidence of putative neural crest cells, which appear as delaminated cells that migrate along pathways corresponding to neural crest cells in fish and amphibians. Molecular cloning studies further revealed the expression of several regulatory genes, including cognates of Pax6, Pax3/7, SoxEa and Sox9, suggesting that the hagfish neural crest is specified by molecular mechanisms that are general to vertebrates. We propose that the neural crest emerged as a population of de-epithelialized migratory cells in a common vertebrate ancestor, and suggest that the possibility of classical and molecular embryology in hagfish opens up new approaches to clarifying the evolutionary history of vertebrates.

Stem Cells & Regenerative Medicine: from Molecular Embryology to Tissue Engineering and Therapeutics

The potential of stem cell research is seemingly unlimited. With this tool, scientists and clinicians are able to focus not just on finding better treatments but also on finding cures for many of the diseases ailing the world today, including heart disease, neurodegenerative diseases, and metabolic diseases such as diabetes. Modern medical science is finally bringing together the intellectual forces of international academic researchers, industry scientists, and clinicians. Such collaborations are of high relevance for emerging science such as stem cell research, which hold great potential for therapeutics and understanding of development and diseases. This sense of collaboration was prominent at the First International Stem Cells & Regenerative Medicine – 2006 Meeting held by GeneExpression Systems in San Francisco, USA in January, which recognized the promise of stem cells and profiled key advancements in various areas of stem cell research. This 3-day meeting brought together industry leaders and entrepreneurs, renowned international scientists from high caliber academic institutions, and even some government representatives, contributing seminars, posters, and product presentations displaying the latest tools in stem cell research. Attendees to this inaugural meeting were greeted with a letter from Governor Arnold Schwarzenegger endorsing the conference, which began with lectures by Nobel Laureate (1980, Chemistry) Dr Paul Berg (Stanford University, Stanford, CA) and by Dr Gail Martin, (University of California, San Francisco, CA, USA). Dr Berg’s presentation covered the saga of pushing stem cell research to the forefront of science, in particular by working with policy makers both at the state and national level in order to gain ‘political justification’. He compared the present stem cell research battle with the mid-1970s recombinant DNA scenario, and is optimistic that stem cell research will be useful for the benefit of human kind, if certain guidelines are followed. Dr Martin covered the historical aspects of stem cell research, speaking from an embryological and developmental biology perspective. She narrated the ‘road to embryonic stem cells and beyond’ beginning with the 1960s and the work of Leroy Stevens from the Jackson labs, who discovered ‘embryonal carcinoma’ cells whilst studying testicular teratocarcinomas. Later Stevens and colleagues demonstrated that these embryonal carcinoma cells are indeed ‘pluripotent stem cells’. In the mid-1970s Dr Martin’s postdoctoral work with Dr Evans at the University of Cambridge, UK led her to develop new and in vitro clonal culture methods of embryoid cells. In the early 1980s she and Evans independently isolated stem cells from mouse embryos and coined the term ‘embryonic stem cells.’ Martin also highlighted the work of Dr Jamie Thompson of the University of Wisconsin, who was successful in culturing the human embryonic stem cell lines during the mid-1990s. Presentations throughout the meeting covered an array of topics staying true to the meeting’s subtitle, Molecular Embryology to Tissue Engineering & Therapeutics.

Evolutionary reorganizations of ontogenesis in ascidians of the genus Molgula

The data on comparative, experimental, and molecular embryology of ascidians (genus Molgula) published during the last 15 years have been reviewed. Some representatives of this genus evolved from development with a tailed larva (tadpole) to direct development associated with the loss of larval structures, such as tail, notochord, sensory organs, and differentiated muscles. The data on evolutionary reorganizations of ontogenesis in ascidians of the genus Molgula have been compared with those in sea urchins, anuran amphibians, and some other organisms.

Mammalian Emi2 mediates cytostatic arrest and transduces the signal for meiotic exit via Cdc20

Fertilizable mammalian oocytes are arrested at the second meiotic metaphase (mII) by the cyclinB-Cdc2 heterodimer, maturation promoting factor (MPF). MPF is stabilized via the activity of an unidentified cytostatic factor (CSF), thereby suspending meiotic progression until fertilization. We here present evidence that a conserved 71 kDa mammalian orthologue of Xenopus XErp1/Emi2, which we term endogenous meiotic inhibitor 2 (Emi2) is an essential CSF component. Depletion in situ of Emi2 by RNA interference elicited precocious meiotic exit in maturing mouse oocytes. Reduction of Emi2 released mature mII oocytes from cytostatic arrest, frequently inducing cytodegeneration. Mos levels autonomously declined to undetectable levels in mII oocytes. Recombinant Emi2 reduced the propensity of mII oocytes to exit meiosis in response to activating stimuli. Emi2 and Cdc20 proteins mutually interact and Cdc20 ablation negated the ability of Emi2 removal to induce metaphase release. Consistent with this, Cdc20 removal prevented parthenogenetic or sperm-induced meiotic exit. These studies show in intact oocytes that the interaction of Emi2 with Cdc20 links activating stimuli to meiotic resumption at fertilization and during parthenogenesis in mammals.

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.

Defining a holoprosencephaly locus on human chromosome 14q13 and characterization of potential candidate genes

Holoprosencephaly (HPE) is the most common developmental field defect in patterning of the human prosencephalon and associated craniofacial structures. The genetics is complex, with 12 loci defined on 11 chromosomes. We defined a locus for HPE (HPE8) on human chromosome 14q13 between markers D14S49 and AFM205XG5, by mapping deletion intervals of affected subjects with proximal chromosome 14q interstitial cytogenetic deletions. A 35-BAC contig was built by chromosome walking. By annotation of the 2.82-Mb minimal critical region, we identified 28 possible genes. Seven genes were expressed in human fetal brain: NPAS3, SNX6, C14ORF11, C14ORF10, PAX9, NKX2.1, and C14ORF19, the last an apparent gene fragment. Molecular embryology, animal modeling, and human mutation studies were reported elsewhere for PAX9 and NKX2.1. We focused on three genes, SNX6, NPAS3, and C14ORF11, as potential candidates for HPE. Genomic structure, human expression patterns, protein cellular localization, and embryonic expression patterns of orthologous murine genes were determined, showing that the three genes have properties similar to those of known HPE genes.

Biochemistry and the Sciences of Recognition

Biochemistry and the Sciences of Recognition

Regulation of outgrowth and apoptosis for the terminal appendage:external genitalia: development by concerted actions of BMP signaling

Extra-corporal fertilization depends on the formation of copulatory organs: the external genitalia. Coordinated growth and differentiation of the genital tubercle (GT), an embryonic anlage of external genitalia, generates a proximodistally elongated structure suitable for copulation, erection, uresis and ejaculation. Despite recent progress in molecular embryology, few attempts have been made to elucidate the molecular developmental processes of external genitalia formation. Bone morphogenetic protein genes (Bmp genes) and their antagonists were spatiotemporally expressed during GT development. Exogenously applied BMP increased apoptosis of GT and inhibited its outgrowth. It has been shown that the distal urethral epithelium (DUE), distal epithelia marked by the Fgf8 expression, may control the initial GT outgrowth. Exogenously applied BMP4 downregulated the expression of Fgf8 and Wnt5a, concomitant with increased apoptosis and decreased cell proliferation of the GT mesenchyme. Furthermore, noggin mutants and Bmpr1a conditional mutant mice displayed hypoplasia and hyperplasia of the external genitalia respectively. noggin mutant mice exhibited downregulation of Wnt5a and Fgf8 expression with decreased cell proliferation. Consistent with such findings, Wnt5a mutant mice displayed GT agenesis with decreased cell proliferation. By contrast, Bmpr1a mutant mice displayed decreased apoptosis and augmented Fgf8 expression in the DUE associated with GT hyperplasia. These results suggest that some of the Bmp genes could negatively affect proximodistally oriented outgrowth of GT with regulatory functions on cell proliferation and apoptosis. The DUE region can be marked only until 14.0 dpc (days post coitum) in mouse development, while GT outgrowth continues thereafter. Possible signaling crosstalk among the whole distal GT regions were also investigated.

Bioelectromagnetics in morphogenesis.

Understanding the factors that allow biological systems to reliably self-assemble consistent, highly complex, four dimensional patterns on many scales is crucial for the biomedicine of cancer, regeneration, and birth defects. The role of chemical signaling factors in controlling embryonic morphogenesis has been a central focus in modern developmental biology. While the role of tensile forces is also beginning to be appreciated, another major aspect of physics remains largely neglected by molecular embryology: electromagnetic fields and radiations. The continued progress of molecular approaches to understanding biological form and function in the post genome era now requires the merging of genetics with functional understanding of biophysics and physiology in vivo. The literature contains much data hinting at an important role for bioelectromagnetic phenomena as a mediator of morphogenetic information in many contexts relevant to embryonic development. This review attempts to highlight briefly some of the most promising (and often underappreciated) findings that are of high relevance for understanding the biophysical factors mediating morphogenetic signals in biological systems. These data originate from contexts including embryonic development, neoplasm, and regeneration.

STEM CELL STANDARDS

APROPOSED SET OF STANdards for evaluating human embryonic stem (hES) cells has been put forth by a group of molecular biologists { Science , 300 , 913, (2003)}. It remains to be seen how their proposal will be received by the wider scientific community and the public. The authors of the proposal explain that their goal was to initiate a scientific discussion that may help to establish standards for evaluating hES cells, in particular, to enable the comparison and classification of existing and future cell lines. Lead coauthor Ali H. Brivanlou, a professor of molecular embryology at Rockefeller University, observes that the number of stem cell lines said to be available to U.S.-funded researchers is anumber that has fluctuated. A lack of standards is the problem with stem cell research today, he explains. We don't know how to appreciate or define stem cells, to assess their qualities. What is a stem cell? As it happened, even before the group could put ...

Congenital and idiopathic scoliosis: clinical and genetic aspects.

Genetic and environmental factors influencing spinal development in lower vertebrates are likely to play a role in the abnormalities associated with human congenital scoliosis (CS) and idiopathic scoliosis (IS). An overview of the molecular embryology of spinal development and the clinical and genetic aspects of CS and IS are presented. Utilizing synteny analysis of the mouse and human genetic databases, likely candidate genes for human CS and IS were identified. Review and synteny analysis. A search of the Mouse Genome Database was performed for "genes," "markers" and "phenotypes" in the categories Neurological and neuromuscular, Skeleton, and Tail and other appendages. The Online Mendelian Inheritance in Man was used to determine whether each mouse locus had a known human homologue. If so, the human homologue was assigned candidate gene status. Linkage maps of the chromosomes carrying loci with possibly relevant phenotypes, but without known human homologues, were examined and regions of documented synteny between the mouse and human genomes were identified. Searching the Mouse Genome Database by phenotypic category yielded 100 mutants of which 66 had been mapped. The descriptions of each of these 66 loci were retrieved to determine which among these included phenotypes of scoliosis, kinky or bent tails, other vertebral abnormalities, or disturbances of axial skeletal development. Forty-five loci of interest remained, and for 27 of these the comparative linkage maps of mouse and human were used to identify human syntenic regions to which plausible candidate genes had been mapped. Synteny analysis of mouse candidate genes for CS and IS holds promise due to the close evolutionary relationship between mice and human beings. With the identification of additional genes in animal model systems that contribute to different stages of spine development, the list of candidate genes for CS and IS will continue to grow.

The Segmented Urbilateria: A Testable Scenario

The idea that the last common ancestor of bilaterian animals (Urbilateria) was segmented has been raised recently on evidence coming from comparative molecular embryology. Leaving aside the complex debate on the value of genetic evidence, the morphological and developmental evidence in favor of a segmented Urbilateria are discussed in the light of the emerging molecular phylogeny of metazoans. Applying a cladistic character optimization procedure to the question of segmentation is vastly complicated by the problem of defining without ambiguity what segmentation is and to what taxa this definition applies. An ancestral segmentation might have undergone many complex derivations in each different phylum, thus rendering the cladistics approaches problematic. Taking the most general definitions of coelom and segmentation however, some remarkably similar patterns are found across the bilaterian tree in the way segments are formed by the posterior addition of mesodermal segments or somites. Postulating that these striking similarities in mesodermal patterns are ancestral, a scenario for the diversification of bilaterians from a metameric ancestor is presented. Several types of evolutionary mechanisms (specialization, tagmosis, progenesis) operating on a segmented ancestral body plan would explain the rapid emergence of body plans during the Cambrian. We finally propose to test this hypothesis by comparing genes involved in mesodermal segmentation.

Ernst haeckel's ontogenetic recapitulation: irritation and incentive from 1866 to our time

Ernst Haeckel (1834-1919) must count among the most widely known biologists of his time. His monographs on radiolarian skeletons, sponges and medusae immediately became standard works, owing partly to lavish illustrations that later on culminated in his "Art Forms in Nature", which markedly influenced the "art nouveau" of the early 20th century. Haeckel's main impact, however, came from his numerous popular books that were crucial in transferring Darwin's ideas to continental Europe. Haeckel's main addition was his claim that ontogeny recapitulates phylogeny. It was partly founded on pre-Darwinian observations by J.F. Meckel and K.E. v. Baer who noticed that vertebrate embryos of different species resemble each other more strongly during early ontogenesis than later on. Wishing to illustrate this clearly, Haeckel clandestinely generalized some figures showing early embryos of animals and Man. This "fraud" provided ammunition for his many adversaries, most of whom felt provoked by his antireligious campaigns. The resulting controversies continued well into the 20th century but then subsided. Quite recently, however, they have flared up again, perhaps in connection with progress in molecular embryology that revealed an amazing evolutionary conservation of genes and their cooperation in signal transduction chains. The scientific publications that triggered this flare, and a selection of "Letters to the Editor" in both international science magazines and the German popular press, serve here to show that Haeckel's idea of recapitulation, while having proven its heuristic value, is still causing considerable irritation. This results from the widespread intuition that the marvels of ontogenesis and other biological phenomena must reflect some "intelligent design" rather than Darwinian evolution.

Making sense of life: explaining biological development with models, metaphors, and machines

Preface Introduction PART ONE Models: Explaining Development without the Help of Genes 1. Synthetic Biology and the Origin of Living Form 2. Morphology as a Science of Mechanical Forces 3. Untimely Births of a Mathematical Biology PART TWO Metaphors: Genes and Developmental Narratives 4. Genes, Gene Action, and Genetic Programs 5. Taming the Cybernetic Metaphor 6. Positioning Positional Information PART THREE Machines: Understanding Development with Computers, Recombinant DNA, and Molecular Imaging 7. The Visual Culture of Molecular Embryology 8. New Roles for Mathematical and Computational Modeling 9. Synthetic Biology Redux-Computer Simulation and Artificial Life Conclusion: Understanding Development Notes References Index

Vascular assembly in natural and engineered tissues.

With the advent of molecular embryology and exploitation of genetic models systems, many genes necessary for normal blood vessel formation during early development have been identified. These genes include soluble effectors and their receptors, as well as components of cell-cell junctions and mediators of cell-matrix interactions. In vitro model systems (2-D and 3-D) to study paracrine and autocrine interactions of vascular cells and their progenitors have also been created. These systems are being combined to study the behavior of genetically altered cells to dissect and define the cellular role(s) of specific genes and gene families in directing the migration, proliferation, and differentiation needed for blood vessel assembly. It is clear that a complex spatial and temporal interplay of signals, including both genetic and environmental, modulates the assembly process. The development of real-time imaging and image analysis will enable us to gain further insights into this process. Collaborative efforts among vascular biologists, biomedical engineers, mathematicians, and physicists will allow us to bridge the gap between understanding vessel assembly in vivo and assembling vessels ex vivo.

Embryology in the medical curriculum.

Embryology as a field is in a period of unprecedented change in its knowledge base. Similarly, this is a period of great change in medical curricular planning. One of the most significant questions in embryology education for medical students is how much of the "new" molecular embryology to mix with the "old" developmental anatomy approach. The other question is the most effective venue for instruction in medical embryology. Not all medical curricula have the same objectives; nor do they use the same educational approach. With that in mind, this review outlines several ways in which medical embryology can be offered and how it can be integrated into the medical curriculum. It also lays out topics that are worthy of inclusion in a modern embryology course or sequence.

A genomic regulatory network for development.

Development of the body plan is controlled by large networks of regulatory genes. A gene regulatory network that controls the specification of endoderm and mesoderm in the sea urchin embryo is summarized here. The network was derived from large-scale perturbation analyses, in combination with computational methodologies, genomic data, cis-regulatory analysis, and molecular embryology. The network contains over 40 genes at present, and each node can be directly verified at the DNA sequence level by cis-regulatory analysis. Its architecture reveals specific and general aspects of development, such as how given cells generate their ordained fates in the embryo and why the process moves inexorably forward in developmental time.

Isolation and community: a review of the role of gap-junctional communication in embryonic patterning.

Gap junctions are specialized channels formed between the membranes of two adjacent cells. They permit the direct passage of small molecules from the cytosol of one cell to that of its neighbor, and thus form a system of cell-cell communication that exists alongside familiar secretion/receptor signaling. Gap junction states can be regulated at many levels by factors such as membrane voltage, pH, phosphorylation state, and biochemical signals. Because of the rich potential for regulation of junctional conductance, and directional and molecular gating (specificity), gap junctional communication (GJC) plays a crucial role in many aspects of normal tissue physiology, as well as in tumor progression. However, arguably the most exciting role for GJC is in the regulation of information flow that takes place during embryonic development. This review summarizes the current knowledge of how GJC controls various aspects of embryonic morphogenesis in both vertebrate and invertebrate systems. Modern molecular embryology approaches have complemented biophysical and ultrastructural data, and we are beginning to unravel the patterning roles of GJC in embryonic events such as the patterning of the embryonic left-right axis, as well as the morphogenesis of the heart and limb. Proteins from the Connexin (Cx) gene family, as well as innexins and ductin, are now beginning to be understood as the basis for GJC underlying important embryonic patterning events.

Unique functions of Sonic hedgehog signaling during external genitalia development

Coordinated growth and differentiation of external genitalia generates a proximodistally elongated structure suitable for copulation and efficient fertilization. The differentiation of external genitalia incorporates a unique process, i.e. the formation of the urethral plate and the urethral tube. Despite significant progress in molecular embryology, few attempts have been made to elucidate the molecular developmental processes for external genitalia. The sonic hedgehog (Shh) gene and its signaling genes have been found to be dynamically expressed during murine external genitalia development. Functional analysis by organ culture revealed that Shh could regulate mesenchymally expressed genes, patched 1 (Ptch1), bone morphogenetic protein 4 (Bmp4), Hoxd13 and fibroblast growth factor 10 (Fgf10), in the anlage: the genital tubercle (GT). Activities of Shh for both GT outgrowth and differentiation were also demonstrated. Shh(-/-) mice displayed complete GT agenesis, which is compatible with such observations. Furthermore, the regulation of apoptosis during GT formation was revealed for the first time. Increased cell death and reduced cell proliferation of the Shh(-/-) mice GT were shown. A search for alterations of Shh downstream gene expression identified a dramatic shift of Bmp4 gene expression from the mesenchyme to the epithelium of the Shh mutant before GT outgrowth. Regulation of mesenchymal Fgf10 gene expression by the epithelial Shh was indicated during late GT development. These results suggest a dual mode of Shh function, first by the regulation of initiating GT outgrowth, and second, by subsequent GT differentiation.