Alpha-dystroglycan is a cell-surface glycoprotein that acts as a receptor for both extracellular matrix proteins containing laminin-G domains and certain arenaviruses. Receptor binding is thought to be mediated by a post-translational modification, and defective binding with laminin underlies a subclass of congenital muscular dystrophy. Here, using mass spectrometry-and NMR-based structural analyses, we identified a phosphorylated O-mannosyl glycan on the mucin-like domain of recombinant alpha-dystroglycan, which was required for laminin binding. We demonstrated that patients with muscle-eye-brain disease and Fukuyama congenital muscular dystrophy, as well as mice with myodystrophy, commonly have defects in a post-phosphoryl modification of this phosphorylated O-linked mannose, and that this modification is mediated by the likeacetylglucosaminyltransferase (LARGE) protein. Our findings expand our understanding of the mechanisms that underlie congenital muscular dystrophy.Diverse post-translational modifications influence the structure and function of many proteins. Dystroglycan (DG) is a membrane protein that requires extensive post-translational processing in order to function as an extracellular matrix receptor. It is comprised of an extracellular α-* To whom correspondence should be addressed. kevin-campbell@uiowa.edu.Supporting Online Material www.sciencemag.org Materials and Methods Figs. S1 to S12 Table S1 NIH Public Access DG subunit and a transmembrane β-DG subunit (1). α-DG serves as a receptor for extracellular matrix laminin G domain-containing ligands such as laminin (1) and agrin (2) in both muscle and brain, and these interactions depend on an unidentified post-translational α-DG modification. α-DG is also the cellular receptor for lymphocytic choriomeningitis virus (LCMV), Lassa fever virus (LFV), and clade C New World arenaviruses (3,4). Although the binding sites for LCMV and LFV on α-DG have not yet been identified, they are thought to overlap with the modification recognized by laminin (5,6).Glycosyltransferase-mediated glycosylation is one form of post-translational modification that can modulate protein structure and function. The main forms in mammals are N-and Oglycosylation, and these are distinguished by how the oligosaccharide moiety links to the amino acid. Mutations in six known or putative glycosyltransferase genes-POMT1 (7), POMT2 (8), POMGnT1 (9), fukutin (10), FKRP (11), and LARGE (12)-have been identified in patients with congenital muscular dystrophy (CMD). These disorders cover a spectrum of abnormalities affecting the brain, eye, and skeletal muscle, and show a dramatic gradient of phenotypic severity ranging from the most devastating in Walker-Warburg syndrome (WWS; OMIM# 236670), to less severe in muscle-eye-brain disease (MEB; OMIM# 253280) and Fukuyama CMD (FCMD; OMIM# 253800), and to mild limb-girdle muscular dystrophies. In these diseases, the ability of α-DG to bind laminin is markedly reduced (13), suggesting that these (putative) glycosyltransferases participa...

Xyloglucans are the principal glycans that interlace cellulose microfibrils in most flowering plants. The mur3 mutant of Arabidopsis contains a severely altered structure of this polysaccharide because of the absence of a conserved ␣ -L -fucosyl-(1 → 2)-␤ -D -galactosyl side chain and excessive galactosylation at an alternative xylose residue. Despite this severe structural alteration, mur3 plants were phenotypically normal and exhibited tensile strength in their inflorescence stems comparable to that of wild-type plants. The MUR3 gene was cloned positionally and shown to encode a xyloglucan galactosyltransferase that acts specifically on the third xylose residue within the XXXG core structure of xyloglucan. MUR3 belongs to a large family of type-II membrane proteins that is evolutionarily conserved among higher plants. The enzyme shows sequence similarities to the glucuronosyltransferase domain of exostosins, a class of animal glycosyltransferases that catalyze the synthesis of heparan sulfate, a glycosaminoglycan with numerous roles in cell differentiation and development. This finding suggests that components of the plant cell wall and of the animal extracellular matrix are synthesized by evolutionarily related enzymes even though the structures of the corresponding polysaccharides are entirely different from each other.

Cell walls of the Arabidopsis mutant mur2 contain less than 2% of the wild-type amount of fucosylated xyloglucan because of a point mutation in the fucosyltransferase AtFUT1. The mur2 mutation eliminates xyloglucan fucosylation in all major plant organs, indicating that Arabidopsis thaliana fucosyltransferase 1 (AtFUT1) accounts for all of the xyloglucan fucosyltransferase activity in Because the mur1 mutation affects several cell wall polysaccharides, whereas the mur2 mutation is specific to xyloglucan, the phenotypes of mur1 plants appear to be caused by structural changes in fucosylated pectic components such as rhamnogalacturonan-II. The normal growth habit and wall strength of mur2 plants casts doubt on hypotheses regarding roles of xyloglucan fucosylation in facilitating xyloglucan-cellulose interactions or in modulating growth regulator activity.