Walker-Warburg syndrome (WWS) is clinically defined as congenital muscular dystrophy accompanied by a variety of brain and eye malformations. It represents the most severe clinical phenotype in a spectrum of alpha-dystroglycan posttranslational processing abnormalities, which share a defect in laminin binding glycan synthesis1. Although six WWS causing genes have been described, only half of all patients can currently be diagnosed genetically2. A cell fusion complementation assay using fibroblasts from undiagnosed WWS individuals identified five novel complementation groups. Further evaluation of one group by linkage analysis and targeted sequencing identified recessive mutations in the isoprenoid synthase domain containing (ISPD) gene. Confirmation of the pathogenicity of the identified ISPD mutations was demonstrated by complementation of fibroblasts with wild-type ISPD. Finally, we show that recessive mutations in ISPD abolish the initial step in laminin binding glycan synthesis by disrupting dystroglycan O-mannosylation. This establishes a novel mechanism for WWS pathophysiology.

Protein O-mannosylation is an essential modification in fungi and animals. Different from most other types of O-glycosylation, protein O-mannosylation is initiated in the endoplasmic reticulum by the transfer of mannose from dolichol monophosphate-activated mannose to serine and threonine residues of secretory proteins. In recent years, it has emerged that even bacteria are capable of O-mannosylation and that the biosynthetic pathway of O-mannosyl glycans is conserved between pro- and eukaryotes. In this review, we summarize the observations that have opened up the field and highlight characteristics of O-mannosylation in the different domains/kingdoms of life.

Protein O mannosylation is a crucial protein modification in uni-and multicellular eukaryotes. In humans, a lack of O-mannosyl glycans causes congenital muscular dystrophies that are associated with brain abnormalities. In yeast, protein O mannosylation is vital; however, it is not known why impaired O mannosylation results in cell death. To address this question, we analyzed the conditionally lethal Saccharomyces cerevisiae protein O-mannosyltransferase pmt2 pmt4⌬ mutant. We found that pmt2 pmt4⌬ cells lyse as small-budded cells in the absence of osmotic stabilization and that treatment with mating pheromone causes pheromone-induced cell death. These phenotypes are partially suppressed by overexpression of upstream elements of the protein kinase C (PKC1) cell integrity pathway, suggesting that the PKC1 pathway is defective in pmt2 pmt4⌬ mutants. Congruently, induction of Mpk1p/Slt2p tyrosine phosphorylation does not occur in pmt2 pmt4⌬ mutants during exposure to mating pheromone or elevated temperature. Detailed analyses of the plasma membrane sensors of the PKC1 pathway revealed that Wsc1p, Wsc2p, and Mid2p are aberrantly processed in pmt mutants. Our data suggest that in yeast, O mannosylation increases the activity of Wsc1p, Wsc2p, and Mid2p by enhancing their stability. Reduced O mannosylation leads to incorrect proteolytic processing of these proteins, which in turn results in impaired activation of the PKC1 pathway and finally causes cell death in the absence of osmotic stabilization.Protein O mannosylation is initiated at the endoplasmic reticulum (ER) by the transfer of mannose from dolichyl phosphate-activated mannose to serine or threonine residues of secretory proteins (52). This reaction is catalyzed by an essential family of protein O-mannosyltransferases (PMTs) that is evolutionarily conserved from yeast to humans (28,52,57). The PMT family is divided into the PMT1, PMT2, and PMT4 subfamilies, whose members include transferases closely related to Saccharomyces cerevisiae Pmt1p, Pmt2p, and Pmt4p, respectively (17,57). In S. cerevisiae the entire PMT family is highly redundant (Pmt1 to Pmt7p), and members of the PMT1 and PMT2 subfamilies show marked similarities and distinctions from PMT4 subfamily members. For example, members of the PMT1 subfamily (Pmt1p and Pmt5p) interact in pairs with members of the PMT2 subfamily (Pmt2p and Pmt3p), whereas the unique representative of the PMT4 subfamily forms homomeric complexes (16). Further, the PMT1/PMT2 and PMT4 subfamilies use different acceptor protein substrates in vivo (10,14).Studies of pmt mutants revealed that protein O mannosylation plays a substantial role in uni-and multicellular eukaryotes. In humans, mutations in POMT1 (protein O-mannosyltransferase 1 gene), which encodes a putative counterpart of the yeast Pmt4p O-mannosyltransferase, result in Walker-Warburg syndrome, which is characterized by severe congenital muscular dystrophy, a neuronal migration defect, and structural abnormalities of the eye (2). Mutations of the Drosophila POMT1 orth...

O-mannosylation is an important protein modification in eukaryotes that is initiated by an evolutionarily conserved family of protein O -mannosyltransferases. The first mammalian protein O -mannosyltransferase gene described was the human POMT1 . Mutations in the hPOMT1 gene are responsible for Walker–Warburg syndrome (WWS), a severe recessive congenital muscular dystrophy associated with defects in neuronal migration that produce complex brain and eye abnormalities. During embryogenesis, the murine Pomt1 gene is prominently expressed in the neural tube, the developing eye, and the mesenchyme. These sites of expression correlate with those in which the main tissue alterations are observed in WWS patients. We have inactivated a Pomt1 allele by gene targeting in embryonic stem cells and produced chimeras transmitting the defect allele to offspring. Although heterozygous mice were viable and fertile, the total absence of Pomt1 – / – pups in the progeny of heterozygous intercrosses indicated that this genotype is embryonic lethal. An analysis of the mutant phenotype revealed that homozygous Pomt1 – / – mice suffer developmental arrest around embryonic day (E) 7.5 and die between E7.5 and E9.5. The Pomt1 – / – embryos present defects in the formation of Reichert's membrane, the first basement membrane to form in the embryo. The failure of this membrane to form appears to be the result of abnormal glycosylation and maturation of dystroglycan that may impair recruitment of laminin, a structural component required for the formation of Reichert's membrane in rodents. The targeted disruption of mPomt1 represents an example of an engineered deletion of a known glycosyltransferase involved in O -mannosyl glycan synthesis.