Biochemical and biophysical changes underlie the mechanisms of basement membrane disruptions in a mouse model of dystroglycanopathy

Abstract

Mutations in glycosyltransferases, such as protein O-mannose N-acetylglucosaminyltransferase 1 (POMGnT1), causes disruptions of basement membranes (BMs) that results in neuronal ectopias and muscular dystrophy. While the mutations diminish dystroglycan-mediated cell-ECM interactions, the cause and mechanism of BM disruptions remain unclear. In this study, we established an in vitro model to measure BM assembly on the surface of neural stem cells. Compared to control cells, the rate of BM assembly on POMGnT1 knockout neural stem cells was significantly reduced. Further, immunofluorescence staining and quantitative proteomic analysis of the inner limiting membrane (ILM), a BM of the retina, revealed that laminin-111 and nidogen-1 were reduced in POMGnT1 knockout mice. Finally, atomic force microscopy showed that the ILM from POMGnT1 knockout mice was thinner with an altered surface topography. The results combined demonstrate that reduced levels of key BM components cause physical changes that weaken the BM in POMGnT1 knockout mice. These changes are caused by a reduced rate of BM assembly during the developmental expansion of the neural tissue.

Human embryoid bodies model basal lamina assembly and muscular dystrophy

“…Our result – that endoderm-free patient embryoid bodies show diminished laminin accumulation – corroborates evidence that αDG-deficient cells have reduced ECM-binding kinetics that might render basement membranes susceptible to mechanically-induced deformation [21, 52]. Crucially, we found that embryoid bodies undergo minimal volumetric growth.…”

“…Knockout of dystroglycan in mouse embryonic stem cells and neural stem cells has been reported to reduce laminin polymerization at the cell surface, which is a prerequisite for basal lamina formation [21, 52]. Because we observed thinner basal lamina in dystroglycanopathy patient embryoid bodies, we next investigated whether this phenotype might be linked to a reduced ability to polymerize laminin.…”

“…A thin and discontinuous basal lamina has been reported in the muscle of dystroglycanopathy patients [25, 48], similar to our observation in patient embryoid bodies. Also, the retina inner limiting basement membrane is thin, patchy, and less stiff in Pomgnt1 -null mice [23, 52]. One caveat of our hiPSC-derived embryoid body system is that it seems to only recapitulate basal lamina development.…”

New concepts in basement membrane biology

“…Mechanically weakened BMs explain the frequent breaks of the ILM and pial BMs that are characteristic for these mice ( Fig. 2A-D) [80,85]. BM ruptures in brain and retina are found in mice and fish with mutations of almost all major BM proteins, including perlecan [28], laminins [30,31,34,35,37,86] and collagen IV [32,36,87].…”

Chitosan facilitates structure formation of the salivary gland by regulating the basement membrane components

“…BM is a dynamic structure whose morphology and composition change in response to development, tissue types, aging, disease, and regeneration [37,38]. We previously reported that de novo BM formation is feasible when the epithelium and mesenchyme of SMGs were cultured in an environment composed of appropriate biomaterials [39], thus proving that the regulation of BM formation and activity is fundamental to promoting epithelial morphogenesis if the developmental scheme is to be recapitulated.…”