Walker-Warburg syndrome (WWS) is an autosomal recessive developmental disorder characterized by congenital muscular dystrophy and complex brain and eye abnormalities. A similar combination of symptoms is presented by two other human diseases, muscle-eye-brain disease (MEB) and Fukuyama congenital muscular dystrophy (FCMD). Although the genes underlying FCMD (Fukutin) and MEB (POMGnT1) have been cloned, loci for WWS have remained elusive. The protein products of POMGnT1 and Fukutin have both been implicated in protein glycosylation. To unravel the genetic basis of WWS, we first performed a genomewide linkage analysis in 10 consanguineous families with WWS. The results indicated the existence of at least three WWS loci. Subsequently, we adopted a candidate-gene approach in combination with homozygosity mapping in 15 consanguineous families with WWS. Candidate genes were selected on the basis of the role of the FCMD and MEB genes. Since POMGnT1 encodes an O-mannoside N-acetylglucosaminyltransferase, we analyzed the possible implication of O-mannosyl glycan synthesis in WWS. Analysis of the locus for O-mannosyltransferase 1 (POMT1) revealed homozygosity in 5 of 15 families. Sequencing of the POMT1 gene revealed mutations in 6 of the 30 unrelated patients with WWS. Of the five mutations identified, two are nonsense mutations, two are frameshift mutations, and one is a missense mutation. Immunohistochemical analysis of muscle from patients with POMT1 mutations corroborated the O-mannosylation defect, as judged by the absence of glycosylation of alpha-dystroglycan. The implication of O-mannosylation in MEB and WWS suggests new lines of study in understanding the molecular basis of neuronal migration.

We report the full cDNA sequence encoding the human homologue of murine PA2.26 (T1␣-2, podoplanin), a small mucin-type transmembrane glycoprotein originally identified as a cell-surface antigen induced in keratinocytes during mouse skin carcinogenesis. The human PA2.26 gene is expressed as 2 transcripts of 0.9 and 2.7 kb in several normal tissues, such as the placenta, skeletal muscle, heart and lung. Using a specific polyclonal antibody raised against a synthetic peptide of the protein ectodomain, PA2.26 was immunohistochemically detected in about 25% (15/61) of human early oral squamous cell carcinomas. PA2.26 distribution in the tumours was heterogeneous and often restricted to the invasive front. Double immunofluorescence and confocal microscopy analysis showed that PA2.26 colocalized with the membrane cytoskeleton linker ezrin at the surface of tumour cells and that its presence in vivo was associated with downregulation of membrane E-cadherin protein expression. Ectopic expression of human PA2.26 in HeLa carcinoma cells and immortalized HaCaT keratinocytes promoted a redistribution of ezrin to the cell edges, the formation of cell-surface protrusions and reduced Ca 2؉ -dependent cell-cell adhesiveness. These results point to PA2.26 as a novel biomarker for oral squamous cell carcinomas that might be involved in migration/invasion.Key words: mucin; PA2.26; ezrin; E-cadherin; microvilli; OSCC Squamous cell carcinomas (SCCs) of the oral cavity, pharynx and larynx remain a significant public health problem. They represent 2-3% of all malignancies, and their incidence, particularly that of oral SCCs (OSCCs), is increasing in Western countries. 1 In spite of improved therapeutic procedures, the prognosis of OSCC patients remains poor and considerably lower than that of other neoplasias. 2 This fact can be attributed to several factors: failure to respond to available chemotherapy, late presentation of the lesions and lack of suitable markers for early detection and prognosis. 3,4 Hence, the finding of novel tumour markers, particularly those associated with tumour cell invasion and spreading, can help provide a more accurate evaluation of prognosis and a more efficient management of the disease.PA2.26 antigen was identified in our laboratory as a cell-surface protein induced in murine epidermal keratinocytes and dermal fibroblast-like cells during wound healing and chemical carcinogenesis. 5 Sequence analysis of the isolated cDNA and biochemical characterization of the protein revealed that murine PA2.26 is a small mucin-like transmembrane glycoprotein of about 45 kDa, 6 highly homologous to the rat alveolar type I cell marker T1␣ and the podocyte-associated glycoprotein podoplanin. 7,8 Murine PA2.26 nucleotide sequence is almost identical to that of OTS-8 and gp38, markers of the osteoblastic cell lineage and stromal cells in peripheral lymphoid tissues, respectively, 9,10 and completely matches the nucleotide sequence of RANDAM-2, a recently discovered membrane glycoprotein expressed in neuronal cells during m...

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.