Reduced expression of EXTL2, a member of the EXT family of glycosyltransferases, in human embryonic kidney 293 cells results in longer heparan sulfate chains [Glycobiology and Extracellular Matrices]

March 31st, 2015 by Katta, K., Imran, T., Busse–Wicher, M., Gronning, M., Czaȷkowski, S., Kusche–Gullberg, M.

Heparan sulfate proteoglycans are ubiquitously located on cell surfaces and in the extracellular matrices. The negatively charged heparan sulfate chains interact with a multitude of different proteins thereby influencing a variety of cellular and developmental processes as for example cell adhesion, migration, tissue morphogenesis and differentiation. The human exostosin (EXT) family of genes contains five members; the heparan sulfate polymerizing enzymes, EXT1 and EXT2, and three EXT-like genes, EXTL1, EXTL2 and EXTL3. EXTL2 has been ascribed activities related to the initiation and termination of heparan sulfate chains. Here we further investigated the role of EXTL2 in heparan sulfate chain elongation by gene-silencing and overexpression strategies. We found that siRNA mediated knockdown of EXTL2 in human embryonic kidney 293 cells resulted in increased chain length, whereas overexpression of EXTL2 in the same cell line had little or no effect on heparan sulfate chain length. To study in more detail the role of EXTL2 in heparan sulfate chain elongation we tested the ability of the overexpressed protein to catalyze the in vitro incorporation of N-acetylglucosamine and N-acetylgalactosamine to oligosaccharide acceptors resembling unmodified heparan sulfate and chondroitin sulfate precursor molecules. Analysis of the generated products revealed that recombinant EXTL2 showed weak ability to transfer N-acetylgalactosamine to heparan sulfate precursor molecules but also, that EXTL2 exhibited much stronger in vitro N-acetylglucosamine transferase activity related to elongation of heparan sulfate chains.
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