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A role for polyglucans in a model sea urchin embryo cellular interaction

Published online by Cambridge University Press:  27 March 2013

Suprita Singh ,
Eddie Karabidian ,
Alexander Kandel ,
Stan Metzenberg ,
Edward J. Carroll Jr  and
Steven B. Oppenheimer
Suprita Singh
Affiliation:
Center for Cancer and Developmental Biology, California State University, 18111 Nordhoff Street, Northridge, California 91330-8303, USA.
Eddie Karabidian
Affiliation:
Center for Cancer and Developmental Biology, California State University, 18111 Nordhoff Street, Northridge, California 91330-8303, USA.
Alexander Kandel
Affiliation:
Center for Cancer and Developmental Biology, California State University, 18111 Nordhoff Street, Northridge, California 91330-8303, USA.
Stan Metzenberg
Affiliation:
Center for Cancer and Developmental Biology, California State University, 18111 Nordhoff Street, Northridge, California 91330-8303, USA.
Edward J. Carroll Jr
Affiliation:
Department of Chemistry and Biochemistry, California State University, 18111 Nordhoff Street, Northridge, California 91330-8262, USA.
Steven B. Oppenheimer*
Affiliation:
Center for Cancer and Developmental Biology, California State University, Northridge, 18111 Nordhoff Street, Northridge, California 91330-8303, USA.
*
All correspondence to: Steven B. Oppenheimer. Center for Cancer and Developmental Biology, California State University, Northridge, 18111 Nordhoff Street, Northridge, California 91330-8303, USA. Tel: +1 818 677 3336. Fax: +1 818 677 2034. e-mail: [email protected]
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Summary

The enzymatic activities of commercially prepared glycosidases were verified by direct chemical assays using defined substrates and fixed and live sea urchin (Lytechinus pictus) embryos to determine if a model cellular interaction of interest to developmental biologists for over a century (interaction of archenteron tip and roof of the blastocoel) was mediated by glycans. Glycosidases (active and denatured) were incubated with microdissected archenterons and blastocoel roofs in a direct assay to learn if their enzymatic activities could prevent the normal adhesive interaction. Of the five glycosidases tested only β-amylase (an exoglycosidase) immediately inhibited the interaction at relatively low unit activity. α-Amylase (an endoglycosidase) had no measurable effect, while other glycosidases (α-glucosidase, β-glucosidase, β-galactosidase) only substantially inhibited adhesion after a 12-h incubation. We demonstrated that the five glycosidases were active (not inhibited) in the presence of embryo materials, and that cleaved sugars could be detected directly after incubation of some enzymes with the embryos. The biochemical purity of the enzymes was examined using gel electrophoresis under denaturing conditions, and the absence of contaminating proteases was confirmed using Azocoll™ substrate. As we cannot entirely rule out the presence of minor contaminating enzymatic activities, only inhibitions of adhesion after very short incubations with enzyme were considered significant and biologically relevant. Although glycans in indirect experiments have been implicated in mediating the interaction of the tip of the archenteron and roof of the blastocoel, to our knowledge, this is the first study that directly implicates polyglucans with terminal 1,4-linked glucose residues in this adhesive event.

Keywords

Archenteron attachmentβ-AmylaseDirect adhesion assayPolyglucansSea urchin embryos
Type
Research Article
Information
Zygote , Volume 22 , Issue 3 , August 2014 , pp. 419 - 429
Copyright
Copyright © Cambridge University Press 2013 

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