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Homodimerization controls the fibroblast growth factor 9 subfamily's receptor binding and heparan sulfate-dependent diffusion in the extracellular matrix.

ABSTRACT: Uncontrolled fibroblast growth factor (FGF) signaling can lead to human diseases, necessitating multiple layers of self-regulatory control mechanisms to keep its activity in check. Herein, we demonstrate that FGF9 and FGF20 ligands undergo a reversible homodimerization, occluding their key receptor binding sites. To test the role of dimerization in ligand autoinhibition, we introduced structure-based mutations into the dimer interfaces of FGF9 and FGF20. The mutations weakened the ability of the ligands to dimerize, effectively increasing the concentrations of monomeric ligands capable of binding and activating their cognate FGF receptor in vitro and in living cells. Interestingly, the monomeric ligands exhibit reduced heparin binding, resulting in their increased radii of heparan sulfate-dependent diffusion and biologic action, as evidenced by the wider dilation area of ex vivo lung cultures in response to implanted mutant FGF9-loaded beads. Hence, our data demonstrate that homodimerization autoregulates FGF9 and FGF20's receptor binding and concentration gradients in the extracellular matrix. Our study is the first to implicate ligand dimerization as an autoregulatory mechanism for growth factor bioactivity and sets the stage for engineering modified FGF9 subfamily ligands, with desired activity for use in both basic and translational research.

SUBMITTER: Kalinina J

PROVIDER: S-EPMC2725704 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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Homodimerization controls the fibroblast growth factor 9 subfamily's receptor binding and heparan sulfate-dependent diffusion in the extracellular matrix.

Kalinina Juliya J Byron Sara A SA Makarenkova Helen P HP Olsen Shaun K SK Eliseenkova Anna V AV Larochelle William J WJ Dhanabal Mohanraj M Blais Steven S Ornitz David M DM Day Loren A LA Neubert Thomas A TA Pollock Pamela M PM Mohammadi Moosa M

Molecular and cellular biology 20090629 17

Uncontrolled fibroblast growth factor (FGF) signaling can lead to human diseases, necessitating multiple layers of self-regulatory control mechanisms to keep its activity in check. Herein, we demonstrate that FGF9 and FGF20 ligands undergo a reversible homodimerization, occluding their key receptor binding sites. To test the role of dimerization in ligand autoinhibition, we introduced structure-based mutations into the dimer interfaces of FGF9 and FGF20. The mutations weakened the ability of the ...[more]

PMID: 19564416

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Project description:The heparan sulfate (HS) chains of proteoglycans are a key regulatory component of the extracellular matrices of animal cells, including the pericellular matrix around the plasma membrane. In these matrices they regulate transport, gradient formation, and effector functions of over 400 proteins central to cell communication. HS from different matrices differs in its selectivity for its protein partners. However, there has been no direct test of how HS in the matrix regulates the transport of its partner proteins. We address this issue by single molecule imaging and tracking in fibroblast pericellular matrix of fibroblast growth factor 2 (FGF2), stoichiometrically labelled with small gold nanoparticles. Transmission electron microscopy and photothermal heterodyne imaging (PHI) show that the spatial distribution of the HS-binding sites for FGF2 in the pericellular matrix is heterogeneous over length scales ranging from 22 nm to several µm. Tracking of individual FGF2 by PHI in the pericellular matrix of living cells demonstrates that they undergo five distinct types of motion. They spend much of their time in confined motion (∼110 nm diameter), but they are not trapped and can escape by simple diffusion, which may be slow, fast, or directed. These substantial translocations (µm) cover distances far greater than the length of a single HS chain. Similar molecular motion persists in fixed cells, where the movement of membrane PGs is impeded. We conclude that FGF2 moves within the pericellular matrix by translocating from one HS-binding site to another. The binding sites on HS chains form non-random, heterogeneous networks. These promote FGF2 confinement or substantial translocation depending on their spatial organisation. We propose that this spatial organisation, coupled to the relative selectivity and the availability of HS-binding sites, determines the transport of FGF2 in matrices. Similar mechanisms are likely to underpin the movement of many other HS-binding effectors.

Project description:UNLABELLED:Nipah virus and Hendra virus are emerging, highly pathogenic, zoonotic paramyxoviruses that belong to the genus Henipavirus. They infect humans as well as numerous mammalian species. Both viruses use ephrin-B2 and -B3 as cell entry receptors, and following initial entry into an organism, they are capable of rapid spread throughout the host. We have previously reported that Nipah virus can use another attachment receptor, different from its entry receptors, to bind to nonpermissive circulating leukocytes, thereby promoting viral dissemination within the host. Here, this attachment molecule was identified as heparan sulfate for both Nipah virus and Hendra virus. Cells devoid of heparan sulfate were not able to mediate henipavirus trans-infection and showed reduced permissivity to infection. Virus pseudotyped with Nipah virus glycoproteins bound heparan sulfate and heparin but no other glycosaminoglycans in a surface plasmon resonance assay. Furthermore, heparin was able to inhibit the interaction of the viruses with the heparan sulfate and to block cell-mediated trans-infection of henipaviruses. Moreover, heparin was shown to bind to ephrin-B3 and to restrain infection of permissive cells in vitro. Consequently, treatment with heparin devoid of anticoagulant activity improved the survival of Nipah virus-infected hamsters. Altogether, these results reveal heparan sulfate as a new attachment receptor for henipaviruses and as a potential therapeutic target for the development of novel approaches against these highly lethal infections. IMPORTANCE:The Henipavirus genus includes two closely related, highly pathogenic paramyxoviruses, Nipah virus and Hendra virus, which cause elevated morbidity and mortality in animals and humans. Pathogenesis of both Nipah virus and Hendra virus infection is poorly understood, and efficient antiviral treatment is still missing. Here, we identified heparan sulfate as a novel attachment receptor used by both viruses to bind host cells. We demonstrate that heparin was able to inhibit the interaction of the viruses with heparan sulfate and to block cell-mediated trans-infection of henipaviruses. Moreover, heparin also bound to the viral entry receptor and thereby restricted infection of permissive cells in vitro. Consequently, heparin treatment improved survival of Nipah virus-infected hamsters. These results uncover an important role of heparan sulfate in henipavirus infection and open novel perspectives for the development of heparan sulfate-targeting therapeutic approaches for these emerging infections.

Dataset Information

Homodimerization controls the fibroblast growth factor 9 subfamily's receptor binding and heparan sulfate-dependent diffusion in the extracellular matrix.

Publications

Homodimerization controls the fibroblast growth factor 9 subfamily's receptor binding and heparan sulfate-dependent diffusion in the extracellular matrix.

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