title: Glycomics of proteoglycan biosynthesis in murine embryonic stem cell differentiation.
abstract: Glycosaminoglycans (GAGs) play a critical role in binding and activation of growth factors involved in cell signaling critical for developmental biology. The biosynthetic pathways for GAGs have been elucidated over the past decade and now analytical methodology makes it possible to determine GAG composition in as few as 10 million cells. A glycomics approach was used to examine GAG content, composition, and the level of transcripts encoding for GAG biosynthetic enzymes as murine embryonic stem cells (mESCs) differentiate to embryoid bodies (EBs) and to extraembryonic endodermal cells (ExE) to better understand the role of GAGs in stem cell differentiation. Hyaluronan synthesis was enhanced by 13- and 24-fold, most likely due to increased expression of hyaluronan synthase-2. Chondroitin sulfate (CS)/dermatan sulfate (DS) synthesis was enhanced by 4- and 6-fold, and heparan sulfate (HS) synthesis was enhanced by 5- and 8-fold following the transition from mESC to EB and ExE. Transcripts associated with the synthesis of the early precursors were largely unaltered, suggesting other factors account for enhanced GAG synthesis. The composition of both CS/DS and HS also changed upon differentiation. Interestingly, CS type E and highly sulfated HS both increase as mESCs differentiate to EBs and ExE. Differentiation was also accompanied by enhanced 2-sulfation in both CS/DS and HS families. Transcript levels for core proteins generally showed increases or remained constant upon mESC differentiation. Finally, transcripts encoding selected enzymes and isoforms, including GlcNAc-4,6-O-sulfotransferase, C5-epimerases, and 3-O-sulfotransferases involved in late GAG biosynthesis, were also enriched. These biosynthetic enzymes are particularly important in introducing GAG fine structure, essential for intercellular communication, cell adhesion, and outside-in signaling. Knowing the changes in GAG fine structure should improve our understanding the biological properties of differentiated stem cells.
authors.affiliation: Complex Carbohydrate Research Center and the University of Georgia, 315 Riverbend Road, Athens, Georgia 30602, USA.
authors.forename: Alison V
authors.initials: AV
authors.lastname: Nairn
authors.forename: Akiko
authors.initials: A
authors.lastname: Kinoshita-Toyoda
authors.forename: Hidenao
authors.initials: H
authors.lastname: Toyoda
authors.forename: Jin
authors.initials: J
authors.lastname: Xie
authors.forename: Kyle
authors.initials: K
authors.lastname: Harris
authors.forename: Stephen
authors.initials: S
authors.lastname: Dalton
authors.forename: Michael
authors.initials: M
authors.lastname: Kulik
authors.forename: J Michael
authors.initials: JM
authors.lastname: Pierce
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Toida
authors.forename: Kelley W
authors.initials: KW
authors.lastname: Moremen
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Linhardt
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D049109:Cell Proliferation; D004187:Disaccharides / Q000737:chemistry; D053595:Embryonic Stem Cells / Q000166:cytology*; D020869:Gene Expression Profiling; D054794:Glycomics / Q000379:methods*; D006025:Glycosaminoglycans / Q000737:chemistry; D051379:Mice; D008954:Models, Biological; D008956:Models, Chemical; D011133:Polysaccharide-Lyases / Q000737:chemistry; D011509:Proteoglycans / Q000737:chemistry*; D012333:RNA, Messenger / Q000378:metabolism; D014212:Tretinoin / Q000737:chemistry
chemical_list: D004187:Disaccharides; D006025:Glycosaminoglycans; D011509:Proteoglycans; D012333:RNA, Messenger; D014212:Tretinoin; D011133:Polysaccharide-Lyases