Heparin IsRequired forCell-Free Binding ofBasic Fibroblast GrowthFactor toaSoluble Receptor and forMitogenesis inWholeCells

Heparin isrequired forthebinding ofbasic fibroblast growth factor (bFGF) tohigh-affinity receptors oncells deficient incell surface heparan sulfate proteoglycan. Sothat this heparin requirement could beevaluated in theabsence ofother cell surface molecules, wedesigned asimple assay based onagenetically engineered soluble formofmurine FGFreceptor 1(mFRl) tagged withplacental alkaline phosphatase. Usingthis assay, we showed thatFGF-receptor binding hasanabsolute requirement forheparin. Byusing acytokine-dependent lymphoid cell line engineered toexpress mFRl,wealso showed that FGF-induced mitogenic activity isheparin dependent. Furthermore, wetested aseries ofsmall heparin oligosaccharides ofdefined lengths fortheir abilities tosupport bFGF-receptor binding andbiologic activity. Wefound that aheparin oligosaccharide with asfewaseight sugar residues issufficient tosupport these activities. We alsodemonstrated thatheparin facilitates FGFdimerization, aproperty thatmaybeimportant forreceptor activation. Heparin orheparan sulfate isrequired forbasic fibroblast growth factor (bFGF) high-affinity receptor binding (38) and forbFGF-induced fibroblast growth andmyoblast differen- tiation (29). Theseobservations suggest that heparan sulfate proteoglycans (HSPGs) maybeimportant regulators of bFGFbiologic activity, acting directly atthelevel ofthecell surface receptor (32). Notsurprisingly, heparin andHSPGs areimportant regulators ofcell growth (37). Depending on thetissue orcell type, heparin caneither stimulate orinhibit cell proliferation (reviewed inreferences 14and31). Someof these effects maybemediated byFGF;however, other growth factors (including granulocyte-macrophage colony- stimulating factor, (GM-CSF), interleukin 3(IL-3), pleiotro- phin, platelet factor 4,keratinocyte autocrine factor or amphiregulin, andheparin-binding-epidermal