Highly stable mutants of human Fibroblast Growth Factor-1 (FGF1)

FGF1 is a powerful mitogen exhibiting strong action on numerous different cell types. This growth factor plays important roles in various stages of development and morphogenesis, as well as in angiogenesis and wound healing processes. FGF1 is a promising candidate for a therapeutic agent, mainly due to its angiogenic, osteogenic and tissue-injury repair properties. Recombinant FGF1 has been shown to induce both rapid angiogenesis and neurogenesis and can find a wide medical application (together with other growth factors) in wound healing. FGF1 has a relatively short half-life in vivo and a denaturation temperature close to physiological. This means that at physiological conditions the protein is almost 50% unfolded and in consequence by many orders of magnitude more sensitive to protease action. Low stability together with sensitivity to protease action questions medical applications of FGF-1. The main aim of our research was to produce and characterize FGF1 variants tailored towards molecular medicine applications. We obtained highly stable variants which not only preserved typical FGF1 properties but also exhibited enhanced mitogenic activity, prolonged in vivo half-life and were more resistant to proteolytic degradation than the wild type1,2,3.




Increased protein stability of FGF1 can compensate for its reduced affinity for heparin

Recently we used highly stable mutants to study the role of heparin in forming the active FGF/FGFR complex. My study showed that increased stability of FGF1 nicely compensates for decreased binding of heparin in FGFR activation and its downstream signaling, induction of DNA synthesis and cell proliferation. Our results indicate that the main role of heparin in FGF-induced signaling is to protect this naturally unstable protein against heat and/or proteolytic degradation and that heparin is not essential for a direct FGF1/FGFR interaction and receptor activation4. This can be of significant medical importance, especially in all cases when addition of heparin to the growth factor is undesirable.

The role of MAP kinases, Erk1 and 2 in direct downregulation of FGFR1 and endocytosis of FGF1-FGFR1 complex

Recently, we have found that upon FGFR1 activation and tyrosine phosphorylation the receptor is directly phosphorylated on Ser777 by activated MAP kinases Erk1 and 2. This serine phosphorylation reduces the tyrosine phosphorylation in the kinase domain of the receptor and provides in this way downregulation mechanism of FGF1 signaling. My experiments reveal a novel MAPK-mediated, negative feedback mechanism that control the signaling via FGFR and protect the cells against unspecific activation of FGFR. We also found that inactivation of Erks reduces rate of FGF1-FGFR1 complex endocytosis.



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