Core O-Glycans Required for Lymphocyte Homing: Gene Knockout Mice of Core 1 β1,3-N-Acetylglucosaminyltransferase and Core 2 N-Acetylglucosaminyltransferase

Abstract Mucin-type O -glycans are synthesized by sequential reaction of glycosyltransferases that have different substrate specificities. To know the significance of specific O -glycan structures, many researchers have been making mice deficient in corresponding enzymes for the synthesis of the O -glycan structures. Here we describe the analysis of gene knockout mice of core 2 branching enzyme (core 2 N -acetylglucosaminyltransferase, Core2GlcNAcT) and core 1 extension enzyme (core 1 β1,3- N -acetylglucosaminyltransferase, Core1-β3GlcNAcT). Because mucin-type O -glycans present sialyl Lewis X (sLeX) and sulfated version of the glycans, which are L-selectin ligands, at the reducing end, the amounts of the ligands of these knockout mice would be reduced. The methods described here are to analyze the interaction between L-selectin and its ligand 6-sulfo sLeX such as lymphocyte homing assay, staining of frozen section, and blotting using L- and E-selectin-IgM chimeric proteins.