O-linked carbohydrates are required for FGF-2-mediated proliferation of mouse embryonic cells

During development, fibroblast growth factors (FGFs) serve highly specific functions that are mediated through high-affinity transmembrane receptors and modulated by membrane-bound proteoglycans. Proteoglycans, in an embryonic environment called embryoglycans, contain numerous carbohydrate ectodomains, the structure of which undergoes rearrangement. Since they can be lost from the cell surface, they are sometimes found in extracellular space where they may also serve some regulatory function. Here we address the potential roles of three naturally occurring isoforms of Lewis X (Le(X)) in FGF-2-mediated proliferation of embryonic stem (ES) cells. We have found that the addition of sulfated Le(X) to ES cells at a concentration of 17 nM promotes FGF-2 mitogenic activity while a 10-fold higher concentration leads to a reduction of FGF-2-mediated proliferation. Notably, this dose-dependent modulation operated only for sulfated Le(X). Other fucosylated motifs, basic Le(X) trisaccharide and sialylated Le(X), also affected ES cell proliferation but the mechanism cannot be clearly correlated with the presence or absence of FGF-2. The suppression of biosynthesis of O-linked carbohydrates including Le(X) reduced basal proliferation of ES cells and interfered with the mitogenic effect of FGF-2. However, in inhibitor-treated cells, the stimulatory activity of FGF-2 can be reestablished to its original level by exogenous Le(X) oligosaccharides. Our results show that (A) O-linked Le(X) oligosaccharides can regulate mitogenic activity of FGF-2 in embryonic cells, (B) and this ability varies with subtle modifications in their structure. Importantly, our data represent the first insight into the mechanism of how growth factor activities might be modulated by shedded embryoglycan ectodomains.