Low fucosylation defines the glycocalyx of progenitor cells and melanocytes in the limbal epithelial stem cell niche
It is widely recognized that the glycocalyx has significant implications in regulating the self-renewal and differentiation of adult stem cells. Yet, its composition remains poorly understood. Here, we show that the fucose-binding Aleuria aurantia lectin (AAL) binds differentially to basal cells in the stratified epithelium of the human limbus, hair follicle and meibomian gland duct. Using fluorescence-activated cell sorting in combination with single-cell transcriptomics, we find that most epithelial progenitor cells and melanocytes in the limbus display low AAL staining (AALlow) on their cell surface, an attribute that is gradually lost in epithelial cells as they differentiate into mature corneal cells. AALlow epithelial cells were enriched in putative limbal stem cell markers and displayed high proliferative potential. Further analyses revealed that AALlow epithelial cells had reduced expression of GDP-mannose-4,6-dehydratase, an enzyme catalyzing the first and regulatory steps in the de novo biosynthesis of GDP-fucose, and that inhibition of fucosylation using a small-molecule fucose analogue stimulated the proliferative potential of limbal epithelial cells ex vivo. These results provide critical insights into the composition of the glycocalyx in adult stem cells and could thereby have implications to the regeneration of the human limbal stem cell niche. Overall design: Single cell transcriptional profiling was performed on AALlow and AALhigh subpopulations to identify the cell types present and the transcript levels of fucosylation-related genes.