Accessing the embryo interior without microinjection

Latham, Virginia H.; Martinez, Anna L.; Cazares, Lisa H.; Hamburger, Howard; Tully, Monica J.; Oppenheimer, Steven B.

doi:10.1016/s0065-1281(98)80027-5

Cited by 17 publications

(41 citation statements)

References 16 publications

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“…Large molecules can enter embryos if added exogenously (Latham et al, 1998) and exert their effects internally as well as externally (Latham et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

Exogenous hyalin and sea urchin gastrulation, Part II: hyalin, an interspecies cell adhesion molecule

Alvarez

Nnoli

Carroll

et al. 2008

Zygote

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SummaryThe 330 kDa fibrillar glycoprotein hyalin is a well known component of the sea urchin embryo extracellular hyaline layer. Only recently, the main component of hyalin, the hyalin repeat domain, has been identified in organisms as widely divergent as bacteria and humans using the GenBank database and therefore its possible function has garnered a great deal of interest. In the sea urchin, hyalin serves as an adhesive substrate in the developing embryo and we have recently shown that exogenously added purified hyalin from Strongylocentrotus purpuratus embryos blocks a model cellular interaction in these embryos, archenteron elongation/attachment to the blastocoel roof. It is important to demonstrate the generality of this result by observing if hyalin from one species of sea urchin blocks archenteron elongation/attachment in another species. Here we show in three repeated experiments, with 30 replicate samples for each condition, that the same concentration of S. purpuratus hyalin (57 μg/ml) that blocked the interaction in living S. purpuratus embryos blocked the same interaction in living Lytechinus pictus embryos. These results correspond with the known crossreactivity of antibody against S. purpuratus hyalin with L. pictus hyalin. We propose that hyalinhyalin receptor binding may mediate this adhesive interaction. The use of a microplate assay that allows precise quantification of developmental effects should help facilitate identification of the function of hyalin in organisms as divergent as bacteria and humans.

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“…Large molecules can enter embryos if added exogenously (Latham et al, 1998) and exert their effects internally as well as externally (Latham et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

Exogenous hyalin and sea urchin gastrulation, Part II: hyalin, an interspecies cell adhesion molecule

Alvarez

Nnoli

Carroll

et al. 2008

Zygote

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“…This is also suggested by our unpublished observations showing that hyalin adheres to mannose /glucose binding lectins in dot blots. Mannose/glucose groups have been implicated in the interaction of the archenteron tip and blastocoel roof as Lens culinaris agglutinin (a mannose/glucose binding lectin) blocked the interaction and was localized, as shown for hyalin, on the blastocoel roof and archenteron surface (Latham et al, 1998;Latham et al, 1999). Also, enzymes that cleave mannose/ glucose residues from glycoproteins, glycolipids and polysaccharides blocked the interaction (Khurrum et al, 2004) and of 22 sugars tested, only a six glucose residue cyclic polysaccharide alpha-cyclodextrin, blocked the interaction (Sajadi, et al, 2007).…”

Section: Discussionmentioning

confidence: 99%

Hyalin is a cell adhesion molecule involved in mediating archenteron–blastocoel roof attachment

et al. 2008

Self Cite

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SummaryThe U. S. National Institutes of Health has designated the sea urchin embryo as a model organism because about twenty-five discoveries in this system have led to insights into the physiology of higher organisms, including humans. Hyalin is a large glycoprotein in the hyaline layer of sea urchin embryos that functions to maintain general adhesive relationships in the developing embryo. It consists of the hyalin repeat domain that has been identified in organisms as diverse as bacteria, worms, flies, mice, sea urchins and humans. Here we show, using a polyclonal antibody raised against the 11.6 S species of hyalin, that it localizes at the tip of the archenteron and on the roof of the blastocoel exactly where these two structures bond in an adhesive interaction that has been of interest for over a century. In addition, the antibody blocks the interaction between the archenteron tip and blastocoel roof. These results, in addition to other recent findings from this laboratory that will be discussed, suggest that hyalin is involved in mediating this cellular interaction. This is the first demonstration that suggests that hyalin is a specific cell adhesion molecule that may function as such in many organisms, including humans.

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“…The developing gut is a simple model for investigating cellular interactions. Because molecules easily enter the interior of sea urchin embryos [6], we are able to precisely quantify the effects of the added reagents on specific cellular interactions. Using these methods we have developed statistically evaluated, quantitative, kinetic graphic profiles of the effects of glycans and glycosidases on the development of the sea urchin archenteron, a model set of cellular interactions, and found that L-rhamnose and polyglucans appear to be involved in these cellular interactions.…”

Section: The Sea Urchin Nih Designated Model System Microplate Assaymentioning

confidence: 99%

Simple Novel Assays in Glycobiology

Oppenheimer

2014

J Glycobiol

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In response to an invitation from this journal, I am providing this mini-review of recent work from the Oppenheimer lab. Over the past 4 decades we have developed many assays that help us examine the role of specifc glycans in cellular interactions. Recently we have used two model systems for this work, the sea urchin embryo and yeast (Saccharomyces cerevisiae). We have developed two assays using the sea urchin embryo. One involves a microplate method where living sea urchin embryos are incubated with specific glycans or glycosidases. A specific set of cellular interactions, development of the primitive gut…archenteron, is examined over time in the presence and absence of the sugars or enzymes in living embryos. L-rhamnose and polyglucans have been identified as playing a role in mediating these cellular interactions. The second assay involves microdissection of the primitive gut away from the blastocoel roof to which it adheres.Using independently characterized glycosidases, we showed that polyglucans appear to mediate this cellular interaction. In the second system using yeast, we examine yeast disaggregation from lectin-derivatized agarose beads in the presence and absence of specific glycans using a quantitative, kinetic graphic profile assay.We found that D-melezitose was the best adhesion inhibitor and may be therapeutically useful in anti-adhesion venues of pathogen binding to cells and in cancer cell binding. The Sea Urchin, NIH Designated Model System Microplate assayWe have used the sea urchin embryo in our glycobiology research for over 4 decades. The sea urchin has been designated by NIH as a model system for understanding mechanisms of importance in human health and disease [1]. Because of its transparency and simplicity, the sea urchin is well suited for research in glycobiology. We have developed new assays using the sea urchin embryo to learn about the roles of specific glycans in mediating cellular interactions [2][3][4][5]. The first assay involves culturing sea urchin embryos in 96 well microplates with and without glycans or glycosidases over time and counting the different morphologies of the developing gut (archenteron) in tens of thousands of living embryos [2,4]. The developing gut is a simple model for investigating cellular interactions. Because molecules easily enter the interior of sea urchin embryos [6], we are able to precisely quantify the effects of the added reagents on specific cellular interactions. Using these methods we have developed statistically evaluated, quantitative, kinetic graphic profiles of the effects of glycans and glycosidases on the development of the sea urchin archenteron, a model set of cellular interactions, and found that L-rhamnose and polyglucans appear to be involved in these cellular interactions. Of importance is that we independently characterize enzymes used in our studies and test for unit activity and contaminants.Assay scheme: Grow sea urchin embryos to 24 hrs or 32 hrs post fertilization in normal artificial sea water. In 96 well micropl...

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Accessing the embryo interior without microinjection

Cited by 17 publications

References 16 publications

Exogenous hyalin and sea urchin gastrulation, Part II: hyalin, an interspecies cell adhesion molecule

Exogenous hyalin and sea urchin gastrulation, Part II: hyalin, an interspecies cell adhesion molecule

Hyalin is a cell adhesion molecule involved in mediating archenteron–blastocoel roof attachment

Simple Novel Assays in Glycobiology

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