Mouse Large Can Modify Complex N- and Mucin O-Glycans on -Dystroglycan to Induce Laminin Binding*

The human LARGE gene encodes a protein with two putative glycosyltransferase domains and is required for the generation of functional -dystroglycan ( -DG). Monoclonal antibodies IIH6 and VIA4-1 recognize the functional glycan epitopes of -DG that are necessary for binding to laminin and other ligands. Overexpression of full-length mouse Large generated functionally glycosylated -DG in Pro 5 Chinese hamster ovary (CHO) cells, and the amount was increased by co-expression of protein:O-mannosyl N-acetylglucosaminyltransferase 1. However, functional -DG represented only a small fraction of the -DG synthesized by CHO cells or expressed from an -DG construct. To identify features of the glycan epitopes induced by Large, the production of functionally glycosylated -DG was investigated in several CHO glycosylation mutants. Mutants with defective transfer of sialic acid (Lec2), galactose (Lec8), or fucose (Lec13) to glycoconjugates, and the Lec15 mutant that cannot synthesize O-mannose glycans, all produced functionally glycosylated -DG upon overexpression of Large. Laminin binding and the -DG glycan epitopes were enhanced in Lec2 and Lec8 cells. In Lec15 cells, functional -DG was increased by co-expression of core 2 N-acetylglucosaminyltransferase 1 with Large. Treatment with N-glycanase markedly reduced functionally glycosylated -DG in Lec2 and Lec8 cells. The combined data provide evidence that Large does not transfer to Gal, Fuc, or sialic acid on -DG nor induce the transfer of these sugars to -DG. In addition, the data suggest that human LARGE may restore functional -DG to muscle cells from patients with defective synthesis of Omannose glycans via the modification of N-glycans and/or mucin O-glycans on -DG.