Conditional knockout of protein O‐mannosyltransferase 2 reveals tissue‐specific roles of O‐mannosyl glycosylation in brain development

Abstract

The meninges produce essential signaling molecules and major protein components of the pial basement membrane during normal brain development. Disruptions in the pial basement membrane underlie neural ectopia seen in those congenital muscular dystrophies (CMDs) caused by mutations in genes involved in O-mannosyl glycosylation. In mammals, biosynthesis of O-mannosyl glycans is initiated by a complex of mutually indispensable protein O-mannosyltransferases 1 and 2 (POMT1 and 2). To study the roles of O-mannosylation in brain development we generated a conditional allele of POMT2. POMT2 nulllizygosity resulted in embryonic lethality because of a defective Reichert's membrane. Brain-specific deletion of POMT2 resulted in hypoglycosylation of α-dystroglycan (DG) and abolished laminin binding activity. The effect of POMT2 deletion on brain development was dependent on timing, as earlier deletion resulted in more severe phenotypes. Multiple brain malformations including overmigration of neocortical neurons and migration failure of granule cells in the cerebellum were observed. Immunofluorescence staining and transmission electron microscopy revealed that these migration defects were closely associated with disruptions in the pial basement membrane. Interestingly, POMT2 deletion in the meninges (and blood vessels) did not disrupt the development of the neocortex. Thus, normal brain development requires protein O-mannosylation activity in neural tissue but not the meninges. These results suggest that gene therapy should be directed to the neural tissue instead of the meninges.

Effect of early embryonic deletion of huntingtin from pyramidal neurons on the development and long-term survival of neurons in cerebral cortex and striatum

“…The specificity and efficacy of these antibodies has been shown previously (Mullen et al, 1992; Wolf et al, 1996; Bloch et al, 2011; Hu et al, 2011). Immunolabeling for Substance P (SP) and for the D1 dopamine receptor was used to study the SP+/D1+ direct pathway striatal neurons and their projections to the globus pallidus internus (GPi) and substantia nigra (SN), while immunolabeling for enkephalin (ENK) was used to study the ENK+ indirect pathway neuron striatal projection system to the globus pallidus externus (GPe).…”

Alterations in sulfated chondroitin glycosaminoglycans following controlled cortical impact injury in mice

“…The immunogen for this antibody is the mouse thymic reticulum. It has been shown to label meningeal fibroblasts in the brain (Hu et al, 2011;Takahashi et al, 2011). In our hands, the antibody detects primarily meninges in the brain and very low expression in the other parts of the brain.…”

Protein O-mannosylation is crucial for E-cadherin–mediated cell adhesion

“…1F) and most likely helped Pomt1 −/− null embryos overcome the bottleneck of early preimplantation lethality observed in our Pomt2 −/− mutants. Such variations in early transcript levels also could explain why lethality of Pomt2 −/− null mice on a different genetic background was not evident until postimplantation development (22). To rule out bias that quantitative differences in Pomt transcript levels might introduce in our functional analyses of O-mannosylation during preimplantation development, we established a method to inhibit POMT enzymatic activity directly in early embryos.…”

“…The most severe cases of O-mannosylation deficiencies, such as Walker-Warburg syndrome and muscle-eye-brain disease, are characterized by neuronal migration disorders that result in fatal brain and eye abnormalities (2). Corresponding animal models of O-mannosylation deficiency show highly disorganized neuroepithelia, which had been attributed to defects in the formation of the glia limitans, a basement membrane that is formed in a α-DG-dependent manner (22,38). However, recent results suggest that these neuronal migration defects are explained only partially by α-DG loss of function.…”