Low Stability of Integrin-Binding Deficient Mutant of FGF1 Restricts Its Biological Activity.

Anna Szlachcic,Daniel Krowarsch,Malgorzata Zakrzewska,Martyna Sochacka,Lukasz Opalinski,Aleksandra Czyrek,Jacek Otlewski

doi:10.3390/cells8080899

Anna Szlachcic, Daniel Krowarsch + Show 5 more

Open Access

https://doi.org/10.3390/cells8080899

Copy DOI

Journal: Cells	Publication Date: Aug 15, 2019
Citations: 9	License type: CC BY 4.0

Affiliation: University of Wrocław

Abstract

Fibroblast growth factor 1 (FGF1) has been shown to interact with integrin αvβ3 through a specific binding site, involving Arg35 residue. The FGF1 mutant (R35E) with impaired integrin binding was found to be defective in its proliferative response, although it was still able to interact with FGF receptors (FGFR) and heparin and induce the activation of downstream signaling pathways. Here, we demonstrate that the lack of mitogenic potential of R35E mutant is directly caused by its decreased thermodynamic stability and susceptibility to proteolytic degradation. Introduction of three stabilizing mutations into R35E variant compensated the effect of destabilizing R35E mutation and restored the proliferation potential of FGF1. Moreover, the stabilized R35E variant regained both anti-apoptotic and wound healing activities, while remaining defective in binding to integrin αvβ3. Our results suggest that the thermodynamic stability and resistance to degradation, rather than the interaction with integrin are required for mitogenic response of FGF1.

Highlights

Fibroblast growth factors (FGFs) form a family of signaling proteins involved in diverse cellular processes such as cell proliferation, migration, angiogenesis, embryonic and fetal development [1,2].FGFs interact with specific transmembrane receptor tyrosine kinases (RTKs), FGF receptors (FGFRs), which include four prevalent types (FGFR1–4) and one lacking an intracellular kinase domain (FGFR5) [3]
Fibroblast growth factor 1 (FGF1) R35E (R50E in the full-length numbering system) variant was reported to be defective in inducing a prolonged FGFR-dependent response, including cell proliferation and sustained downstream signaling [6,7]. This was perceived as a result of its defective integrin αv β3 binding, suggesting the importance of ternary FGF1/FGFR/integrin αv β3 complex formation for FGF1-induced signaling
We found that both R35E mutants efficiently bound FGF receptors on the cell surface, as they could outcompete binding of fluorescently-labeled wild-type FGF1 in a concentration-dependent manner (Figure 3A, Figure S1A)

Summary

Introduction

Fibroblast growth factors (FGFs) form a family of signaling proteins involved in diverse cellular processes such as cell proliferation, migration, angiogenesis, embryonic and fetal development [1,2]. FGFs interact with specific transmembrane receptor tyrosine kinases (RTKs), FGF receptors (FGFRs), which include four prevalent types (FGFR1–4) and one lacking an intracellular kinase domain (FGFR5) [3]. FGF-FGFR interaction stimulates receptor dimerization and activation of receptor tyrosine kinases, initiating multiple signal transduction pathways. Regulatory mechanisms of FGFR signaling are not yet fully understood, a growing number of proteins affecting the FGF ligands and FGF-induced signaling cascades have been identified [2]. One of them is integrin αv β3 reported to bind directly to fibroblast growth factor 1

Methods

Results

Conclusion