Specific Molecular Interactions of Oversulfated Chondroitin Sulfate E with Various Heparin-binding Growth Factors

Abstract

We previously observed that the cortical neuronal cell adhesion mediated by midkine (MK), a heparin (Hep)-binding growth factor, is specifically inhibited by oversulfated chondroitin sulfate-E (CS-E) (Ueoka, C., Kaneda, N., Okazaki, I., Nadanaka, S., Muramatsu, T., and Sugahara, K. Proteoglycans (PGs)1 that bear heparan sulfate (HS) glycosaminoglycan side chains have attracted much attention because of the demonstration of the involvement of HS in the developmental processes and specific signaling pathways (for review, see Refs. 1-3). Although the chondroitin sulfate (CS) glycosaminoglycan side chains of CS-PGs have attracted less attention until recently, accumulating evidence suggests the importance of these molecules in various biological functions (2, 4). CS-PGs are also ubiquitous components of the extracellular matrix of connective tissues and are also found at the surfaces of many cell types and in intracellular secretory granules (for review, see Refs. 5 and 6). Developmentally regulated expression of CS epitopes in the rodent fetus has been demonstrated by immunological studies (7), especially during central nervous system development (8 -10). CS is a rich component in the extracellular matrix of the brain (11). Developmentally regulated expression and tissue-specific distribution of CS variants suggest that CS chains differing in the degree and profile of sulfation perform distinct functions during development (12).Among variant forms of CS chains, oversulfated CS chains, such as CS-E and CS-D, are of special interest. The presence of E (GlcUA␤1-3GalNAc(4S,6S)) and D (GlcUA(2S)␤1-3GalNAc(6S)) units in bovine brain (13) and E18 rat brain (14) has been reported (GlcUA stands for D-glucuronic acid whereas 2S, 4S, and 6S represent 2-O-, 4-O-, and 6-O-sulfate, respectively). These units are present in appreciable proportions in chick brains, and their expression is regulated developmentally (15). Faissner et al. (16) demonstrated that the neurite outgrowth-stimulating capacity of DSD-1-PG, derived from neonatal mouse brains, is strongly reduced by the monoclonal antibody 473HD, which recognizes a CS epitope, and the interaction is inhibited by shark cartilage CS-C. We have shown that CS-D, an oversulfated CS also derived from shark cartilage, inhibits the interactions between the monoclonal antibody 473HD and DSD-1-PG and also promotes neurite outgrowth of E18 hippocampal neurons (17,18) and that the DSD-1-PG contains the D disaccharide unit, which is a rich and poor component in CS-D and CS-C, respectively. We have further observed that another oversulfated variant CS-E, derived from squid cartilage, also exhibits neurite outgrowth promoting activity, which is not inhibited by 473HD, suggesting a

Sulfation patterns of exogenous chondroitin sulfate affect chondrogenic differentiation of ATDC5 cells

“…Among the CS variants, CS-E and CS-D, which are highly sulfated, promote the outgrowth of neurites in embryonic mouse hippocampal neurons [23]. These two CS variants also show strong affinity to some growth factors, such as fibroblast growth factors and bone morphogenetic proteins (BMPs) [24,25]. CS-E enhances osteoblast differentiation by binding to BMP-4, which is an essential growth factor for chondrogenic differentiation [26].…”

Structural and Functional Characterization of Oversulfated Chondroitin Sulfate/Dermatan Sulfate Hybrid Chains from the Notochord of Hagfish

“…The binding of growth factors (including FGF-2) to HS and DS requires IdoUA residues (43,44). However, CS-E exclusive of IdoUA also exhibits specific interactions of high affinity with various heparin-binding growth factors (13), implying the importance of the E unit and the sulfation patterns as well. CS/DS hybrid chains from embryonic pig brain bind various growth factors (16), and hybrid chains of endocan DS-PG on the endothelial surface are also instrumental in bringing about hepatocyte growth factor-me- g as uronic acid) were digested extensively with chondroitinase ABC (A), AC-I (B), or B (C), and the individual digests were labeled with the fluorophore 2-AB and analyzed by anion-exchange HPLC on a PA-03 column using a 16 -530 mM NaH 2 PO 4 gradient over a 1-h period.…”

“…Digestions with various chondroitinases revealed the involvement of both GlcUA and IdoUA. CS/DS chains bind heparin-binding growth factors (13,16) and may act as receptors or coreceptors in addition to conventional HS chains, and such growth factors may, in turn, be involved in the neuritogenic mechanism of CS/DS chains in vivo. Indeed, Bao et al 2 recently identified PTN as an endogenous ligand for embryonic pig brain CS/DS chains and observed stimulation of the neuritogenic activity of these chains by exogenously added PTN.…”

“…We and others have shown the importance of this class of molecule, from simple chondroitin involved in cell division of a nematode (9) to differentially oversulfated CS-D and CS-E involved in neuroregulatory functions (10 -12) and the binding of growth factors in mammalian systems (13). While pursuing the critical structural elements in the CS variants, we became * This work was supported in part by the Science Research Promotion Fund of the Japan Private School Promotion Foundation and Grant-inaid for Exploratory Research 15659021 and the National Project on Functional Glycoconjugate Research Aimed at Developing New Industry of the Ministry of Education, Science, Sports, and Culture of Japan.…”

Carbohydrate post-glycosylational modifications

“…For example, the squid cartilage CS-E [GlcAβ1,3GalNAc(4S,6S)β1,4] has been found to interact with various heparin-binding growth factors. 49 CS octa or larger oligosaccharides with at least three continuous CS-E sequences have been shown to bind to type V collagen. 50 Purified squid N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase (GalNAc4S-6ST) has been used in enzymatic synthesis of CS-E from CS-A [GlcAβ1,3GalNAc(4S)β1,4], converting half of GalNAc(4SO 4 ) residues to GalNAc(4,6-SO 4 ) residues.51 This enzyme can also catalyze the synthesis of oversulfated DS IdoAα1,3GalNAc(4S,6S)β1,4 units when DS IdoAα1,3GalNAc(4S)β1,4 is used as the substrate.…”