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 a nd modulated by membrane-bound proteoglycans. Proteoglycans, in an embryoni c environment called embryoglycans, contain numerous carbohydrate ectodomai ns, the structure of which undergoes rearrangement. Since they can be lost from the cell surface, they are sometimes found in extracellular space wher e they may also serve some regulatory function. Here we address the potenti al 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 promote s FGF-2 mitogenic activity while a 10-fold higher concentration leads to a reduction of FGF-2-mediated proliferation. Notably, this dose-dependent mod ulation operated only for sulfated Le(X). Other fucosylated motifs, basic L e(X) trisaccharide and sialylated Le(X), also affected ES cell proliferatio n but the mechanism cannot be clearly correlated with the presence or absen ce of FGF-2. The suppression of biosynthesis of O-linked carbohydrates incl uding Le(X) reduced basal proliferation of ES cells and interfered with the mitogenic effect of FGF-2. However, in inhibitor-treated cells, the stimul atory activity of FGF-2 can be reestablished to its original level by exoge nous Le(X) oligosaccharides. Our results show that (A) O-linked Le(X) oligo saccharides can regulate mitogenic activity of FGF-2 in embryonic cells, (B ) and this ability varies with subtle modifications in their structure. Imp ortantly, our data represent the first insight into the mechanism of how gr owth factor activities might be modulated by shedded embryoglycan ectodomai ns.