N-glycosylation Branching and Neural Stem Cell Fate Potential
Neural stem/progenitor cells (NSPCs) generate all the differentiated cell types in the central nervous system. However, mechanisms controlling differentiation into specific cell types, such as neurons and astrocytes, and how differentiation is regulated by extracellular cues must be identified. Gaining a better understanding of how NSPC differentiation is affected by their environment will be essential for developing these cells as therapeutics for brain repair. Our lab identified a novel biophysical signature, membrane capacitance, that indicates neuron- or astrocyte-bias of NSPCs. I found that N-glycosylation, specifically the N-glycan branching pathway, differs between astrocyte- and neuron-biased NSPCs. My studies showed that N-acetylglucosamine (GlcNAc) treatment to enhance N-glycan branching shifted membrane capacitance and altered fate of NSPCs. I identified the MGAT5 branching enzyme as a critical regulator of NSPC differentiation in vitro and in vivo. My studies showed that loss of MGAT5 significantly impacted neural development, causing enhanced neuron differentiation but depletion of the NSPC pool that led to a reduction in neurons in the MGAT5 cortex in vivo. I found that MGAT5 null NSPCs in vitro formed more neurons and fewer astrocytes than their WT counterparts. To identify a potential mechanism for how branching impacts fate, I used a proteomic screen to identify cell surface proteins regulated by branching and found a reduction in cell adhesion proteins in GlcNAc-treated NSPCs. My functional assays probing cell-cell and cell-ECM adhesion showed that cellular adhesion is diminished when NSPC N-glycan branching is enhanced. Taken together, my studies identified N-glycan branching as a critical component of fate-specific membrane capacitance, a novel regulator of NSPC differentiation, and a means by which NSPC differentiation can be affected by the extracellular environment.