AbstractGlycosaminoglycans are sulfated polysaccharide molecules, essential for many biological processes. The 6-O sulfation of glycosaminoglycans is carried out by carbohydrate 6-O sulfotransferases (C6OST), previously named Gal/GalNAc/GlcNAc 6-O sulfotransferases. Here for the first time we present a detailed phylogenetic reconstruction, analysis of gene synteny conservation and propose an evolutionary scenario for the C6OST family in major vertebrate groups, including mammals, birds,

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... n reptiles, amphibians, lobe-finned fishes, ray-finned fishes, cartilaginous fishes and jawless vertebrates. The C6OST gene expansion likely occurred in the chordate lineage, after the divergence of tunicates and before the emergence of extant vertebrate lineages.The two rounds of whole genome duplication in early vertebrate evolution (1R/2R) only contributed two additional C6OST subtype genes, increasing the vertebrate repertoire from four genes to six, divided into two branches. The first branch includes CHST1 and CHST3 as well as a previously unrecognized subtype, CHST16, that was lost in amniotes. The second branch includes CHST2, CHST7 and CHST5. Subsequently, local duplications of CHST5 gave rise to CHST4 in the ancestor of tetrapods, and to CHST6 in the ancestor of primates.The teleost specific gene duplicates were identified for CHST1, CHST2 and CHST3 and are result of whole genome duplication (3R) in the teleost lineage. We could also detect multiple, more recent lineage-specific duplicates. Thus, the repertoire of C6OST genes in vertebrate species has been shaped by different events at several stages during vertebrate evolution, with implications for the evolution of the skeleton, nervous system and cell-cell interactions.