Abstract
Chondroitin/dermatan sulfate (CS/DS) proteoglycans consist of unbranched sulfated polysaccharide chains of repeating GalNAc-GlcA/IdoA disaccharide units, attached to serine residues on specific proteins. The CS/DS proteoglycans are abundant in the extracellular matrix where they have essential functions in tissue development and homeostasis. In this report a phylogenetic analysis of vertebrate genes coding for the enzymes that modify CS/DS is presented. We identify single orthologous genes in the zebrafish genome for the sulfotransferases chst7, chst11, chst13, chst14, chst15 and ust and the epimerase dse. In contrast, two copies were found for mammalian sulfotransferases CHST3 and CHST12 and the epimerase DSEL, named chst3a and chst3b, chst12a and chst12b, dsela and dselb, respectively. Expression of CS/DS modification enzymes is spatially and temporally regulated with a large variation between different genes. We found that CS/DS 4-O-sulfotransferases and 6-O-sulfotransferases as well as CS/DS epimerases show a strong and partly overlapping expression, whereas the expression is restricted for enzymes with ability to synthesize di-sulfated disaccharides. A structural analysis further showed that CS/DS sulfation increases during embryonic development mainly due to synthesis of 4-O-sulfated GalNAc while the proportion of 6-O-sulfated GalNAc increases in later developmental stages. Di-sulfated GalNAc synthesized by Chst15 and 2-O-sulfated GlcA/IdoA synthesized by Ust are rare, in accordance with the restricted expression of these enzymes. We also compared CS/DS composition with that of heparan sulfate (HS). Notably, CS/DS biosynthesis in early zebrafish development is more dynamic than HS biosynthesis. Furthermore, HS contains disaccharides with more than one sulfate group, which are virtually absent in CS/DS.
Highlights
Chondroitin/dermatan sulfate (CS/DS) proteoglycans consist of core proteins with attached linear sulfated polysaccharides of repeating glucuronic acid and N-acetylgalacotsamine (GlcA-GalNAc) in CS or iduronic acid and N-acetylgalactosamine (IdoA-GalNAc) in DS (Fig. 1)
Most CS/DS modification enzymes are present as single orthologues in zebrafish
Amino acid sequence alignments of identified zebrafish enzymes and human orthologues can be found in S1 Fig. We included another teleost fish, stickleback (G. aculeatus), into the analysis in order to better understand the divergence of genes in different teleost lineages after the teleost specific genome duplication
Summary
Chondroitin/dermatan sulfate (CS/DS) proteoglycans consist of core proteins with attached linear sulfated polysaccharides of repeating glucuronic acid and N-acetylgalacotsamine (GlcA-GalNAc) in CS or iduronic acid and N-acetylgalactosamine (IdoA-GalNAc) in DS (Fig. 1). CS/DS Biosynthesis in Zebrafish doi:10.1371/journal.pone.0121957.g001 deposited extensively in cartilage ECM, where the high overall charge provides electrostatic forces to promote high water content, enabling joints to withstand mechanical compression. The biosynthesis of CS/DS is initiated with the synthesis of a GlcA–Gal–Gal–Xyl–O-Ser tetrasaccharide structure attached to one of more than 30 known CS/DS core proteins [5, 6]. This structure constitutes the linkage region for CS/DS as well as for heparan sulfate (HS). In HS biosynthesis, a GlcNAc is added to the linkage region as the fifth sugar followed by repeating additions of GlcNAc and GlcA. Elongation of CS/DS by repeating additions of GalNAc and GlcA is the result of a concerted action of CS/DS glycosyltransferases [5, 6, 10]
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