Cellular signaling by fibroblast growth factor receptors

Cytokine & Growth Factor Reviews 16 (2005) 139–149 www.elsevier.com/locate/cytogfr

Cellular signaling by fibroblast growth factor receptors V.P. Eswarakumar, I. Lax, J. Schlessinger* Yale University School of Medicine, Department of Pharmacology, 333 Cedar Street, P.O. Box 208066, SHM B-295, New Haven, CT 06520, USA Available online 1 February 2005

Abstract The 22 members of the fibroblast growth factor (FGF) family of growth factors mediate their cellular responses by binding to and activating the different isoforms encoded by the four receptor tyrosine kinases (RTKs) designated FGFR1, FGFR2, FGFR3 and FGFR4. Unlike other growth factors, FGFs act in concert with heparin or heparan sulfate proteoglycan (HSPG) to activate FGFRs and to induce the pleiotropic responses that lead to the variety of cellular responses induced by this large family of growth factors. A variety of human skeletal dysplasias have been linked to specific point mutations in FGFR1, FGFR2 and FGFR3 leading to severe impairment in cranial, digital and skeletal development. Gain of function mutations in FGFRs were also identified in a variety of human cancers such as myeloproliferative syndromes, lymphomas, prostate and breast cancers as well as other malignant diseases. The binding of FGF and HSPG to the extracellular ligand domain of FGFR induces receptor dimerization, activation and autophosphorylation of multiple tyrosine residues in the cytoplasmic domain of the receptor molecule. A variety of signaling proteins are phosphorylated in response to FGF stimulation including Shc, phospholipase-Cg, STAT1, Gab1 and FRS2a leading to stimulation of intracellular signaling pathways that control cell proliferation, cell differentiation, cell migration, cell survival and cell shape. The docking proteins FRS2a and FRS2b are major mediators of the Ras/MAPK and PI-3 kinase/Akt signaling pathways as well as negative feedback mechanisms that fine-tune the signal that is initiated at the cell surface following FGFR stimulation. # 2004 Elsevier Ltd. All rights reserved. Keywords: Cellular signaling; FGFR stimulation; Dimerization

1. Introduction The first fibroblast growth factor (FGF) was discovered as a mitogen for cultured fibroblasts [1]. Since then, at least 22 distinct FGFs have been identified in a variety of organisms from nematode and drosophila to mouse and human (reviewed in [2]). Although, FGFs vary in size from 17 to 34 kDa, all members of the family share a conserved sequence of 120 amino acids that show 16–65% sequence identity (reviewed in [2]). FGFs mediate a variety of cellular responses during embryonic development and in the adult organism. During embryonic development, FGFs play a critical role in morphogenesis by regulating cell proliferation, differentiation and cell migration. In the adult organism, FGFs play an important role in the control of the nervous system, in tissue repair, wound healing and in tumor * Corresponding author. Tel.: +1 203 785 7345. E-mail address: [email protected] (J. Schlessinger). 1359-6101/$ – see front matter # 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.cytogfr.2005.01.001

angiogenesis (reviewed in [3]). FGFs mediate their cellular responses by binding to and activating a family of four receptor tyrosine kinases (RTKs) [4–6] designated the highaffinity FGF-receptors FGFR1–FGFR4. FGFs also bind to heparin or heparan sulfate proteoglycans (HSPG), lowaffinity receptor that do not transmit a biological signal but rather function as an accessory molecule that regulate FGFbinding and the activation of the occupied signaling receptors [7–11]. Like all receptor tyrosine kinases, the four signaling FGFR1–FGFR4 are composed of an extracellular ligandbinding domain, a single transmembrane domain and a cytoplasmic domain containing the catalytic protein tyrosine kinase core as well as additional regulatory sequences [12,13]. The extracellular ligand-binding domain of FGFR is composed of three immunoglobulin (Ig) like domains, designated D1–D3; a stretch of seven to eight acidic residues in the linker connecting D1 and D2, designated the ‘‘acid box’’ and a conserved positively charged region in D2 that serves as a binding site for heparin [14].

140

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

2. Isoforms of FGFR

Table 1 Ligand specificities of FGFR isoforms

An important hallmark of the FGFR family of RTKs is that a variety of FGFR isoforms are generated by alternative splicing of FGFR transcripts. The different FGFR isoforms include FGFR with an extracellular domain composed of either two or three Ig-like domains, soluble secreted FGFR forms as well as alternative splicing in the third Ig-like domain (D3) that profoundly alters ligand-binding specificity [15,16]. The alternative splicing in D3 exists in FGFR1, 2 and 3, but not in FGFR4. It has been shown that exon 7 of FGFR2 gene encodes for the N-terminal half of D3 (designated ‘a’), while exons 8 and 9 alternatively encode for the C-terminal half of D3 and are thus designated as ‘b’ and ‘c’ forms of FGFR, respectively (Fig. 1). The two alternative forms display different ligand-binding characteristics. For example, while FGFR2b binds FGF7 and FGF10, but not FGF2, the FGFR2c isoform binds FGF2 and FGF18, but not FGF7 and FGF10. Table 1 summarizes the specificity of these isoforms towards different FGF ligands. Furthermore, it has been shown that the FGFR2b isoform is exclusively expressed in epithelial cells (also designated as KGFR), and that the FGFR2c is expressed exclusively in mesenchymal cells [17]. The lineage-specific expression of the IIIb and IIIc isoforms of FGFRs enables interaction between the epithelial and mesenchymal layers during development in response to different FGFs.

FGFR isoform

Ligand specificity

FGFR1b FGFR1c FGFR2b FGFR2c FGFR3b FGFR3c FGFR4

FGF1, -2, -3 and -10 FGF1, -2, -4, -5 and -6 FGF1, -3, -7, -10 and -22 FGF1, -2, -4, -6, -9, -17 and -18 FGF1 and -9 FGF1, -2, -4, -8, -9, -17, -18 and -23 FGF1, -2, -4, -6, -8, -9, -16, -17, -18 and -19

3. Structure of FGF-receptors In order to reveal the molecular mechanism underlying FGF- and heparin-induced FGFR dimerization and activation, the crystal structures of the ligand-binding domains of FGFR1 and FGFR2 in complex with FGF1 or FGF2, as well as the crystal structure of a ternary FGF/heparin/FGFR

Adapted from [52] and [53]. FGF22 and FGF23 specificities are according to [54] and [55], respectively. FGF7 and 10 are considered to be mesenchymally expressed, and FGF 2, 4, 6, 8, 9 and 17 are considered to be expressed in epithelia.

complex were determined [14,18,19], (Fig. 2A). On the basis of the structural analyses and earlier biochemical studies, it is possible to propose a model for how ligandbinding induces the dimerization and activation of FGFR. The X-ray structures have shown that ligand-bound activated dimeric FGFR is stabilized by both FGF-mediated and direct receptor–receptor interactions as well as by binding of heparin to a positively charged crevice created by the D2 domains of the two FGFRs in the dimer and in the two adjoining bound FGF molecule [14] (Fig. 2). Binding experiments with FGFs and heparin have demonstrated that the binding affinity of both ligands is enhanced towards cells expressing FGFR deletion mutants devoid of D1 and the acid box as compared to their binding to cells expressing intact FGFR [20]. A similar conclusion was reached from binding experiments in which plasmon resonance measurements were applied to determine the kinetic parameters and dissociation constants of FGF1 and heparin towards intact soluble FGFR3c extracellular domain (D1, D2, D3) as compared to an FGFR3c deletion mutant devoid of D1 and the acid box [21]. Moreover, surface plasmon resonance binding experiments have shown that an isolated fragment of

Fig. 1. FGFR isoforms generated by alternative splicing of FGFR transcripts. The two forms of FGFR are generated by alternative splicing of exons 8 and 9. The C-terminal half o f DIII is encoded by exon 8 to generate the FGFR-IIIb isoform while the C-terminal half of DIII is encoded by exon 9 to generate the FGFR-IIIc isoform.

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

141

Fig. 2. Crystal structures of a ternary FGF2/heparin/FGFR1 complex and the catalytic PTK domain of FGFR1:(A) Ribbon diagram of the ternary FGF2/ heparin/FGFR1 complex showing FGF2 in yellow, D2 and D3 domains of the ligand-binding portion in green and blue, and heparin in red. (B) Ribbon diagram of the PTK domain of FGFR1 showing a-helices in red, b-strands in green, nucleotide-binding loop in orange, catalytic loop in blue and activation loop in purple. The side chains of tyr653 and 654 are in yellow. For additional details, see references [23–30].

FGFR3c composed of D1 and an acid box binds specifically to the ligand-binding portion of FGFR3c (D2–D3). In addition, analysis of the crystal structure of the intact extracellular domain of FGFR3c in complex with FGF1 have shown that D1 and the region connecting D1 to the rest of the receptor molecule are disordered while the structure of the ligand-binding portion of FGFR3c (D2–D3) in complex with FGF1 is very similar to the structures of FGF1 or FGF2 in complex with deletion mutants of FGFR1 and FGFR2 comprising only the ligand-binding portions (D2–D3). These experiments provide further support for the assignment of D2 and D3 domain of FGFRs as the primary binding pocket for FGFs and demonstrate that D1 and the acid box has an autoinhibitory function. On the basis of these experiments, we have proposed that a autoinhibited ‘‘closed’’ configuration of FGFR exists in equilibrium with an active ‘‘open’’ state poised towards dimerization, transautophosphorylation and stimulation of PTK activity [21]. The preferred binding of FGF and the accessory molecule heparin to the open, dimerization-competent configuration will shift the equilibrium towards the dimeric form of FGFR resulting in autophosphorylation and stimulation of PTK activity [22]. We have also determined the crystal structure of the protein tyrosine kinase domain of FGFR1 in complex with either an ATP analogue or in complex with a variety of protein tyrosine kinase inhibitors [23–25]. The crystal

structure of the PTK domain of FGFR provides a view of how the activation loop in the catalytic domain of FGFR maintains the PTK domain in an inactive or low activity state [23]. The activation-loop of FGFR1 contains two tyrosine residues that must be autophosphorylated for maintaining the catalytic domain in an active state. In the autoinhibited configuration, a kinase invariant proline residue at the Cterminal end of the activation loop interferes with substrate binding while allowing access to ATP in the nucleotidebinding site (Fig. 2B). The PTK domain of FGFR1 is, thus, less tightly autoinhibited than the PTK domain of the insulin receptor in which the activation loop prevents both substrate and ATP binding to the catalytic core [23]. We have also determined the crystal structure of the PTK domain of FGFR1 in complex with a variety of inhibitors belonging to two different chemical classes: oxindoles and pyridopyrimidines [24,25]. Both the oxindole and the pyridopyrimidine compounds bind specifically to the nucleotidebinding sites in the PTK domain of FGFR1. However, the structural analyses showed that the exact mechanism of inhibition for each compound differs. While the oxindole form a specific hydrogen bond with a key residue in the catalytic domain resulting in a conformational change in the catalytic loop [24], the higher affinity and selectivity of the pyrido-pyrimidines derives from the greater surface complementarity with the ATP binding cleft [25]. On the basis of this detailed structural information, new inhibitors have

142

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

been synthesized combining the favorable features of both the oxindoles and pyrido-pyrimidines. Moreover, the information obtained from the structural studies was used to design more potent inhibitors. Some of these inhibitors are currently being tested in clinical trials for the treatment of various human cancers [26].

immediately after birth. Disruption of the FGFR2c isoform, on the other hand, results in impairment in skull and bone development, but the mutant mice are viable. Finally, disruption of the FGFR3 gene results in bone overgrowth and no obvious phenotype was observed in FGFR4
/
mice.

4. Biological roles of FGF and FGFR isoforms

5. Cell signaling via FGF-receptors

The biological roles of more than half of the 22 known mammalian FGFs have been investigated by targeting the genes of individual FGFs by homologous recombination. The results presented in Table 2 summarizes the phenotypes caused by targeted disruption of 15 out of the 22 FGFs [56–73]. These studies together with analyses of the role played by FGFs in specific developmental systems and the analyses of the expression patterns of FGFs in different tissues and cells, demonstrate that FGFs play critical roles during most stages of mouse development and organogenesis [2]. Furthermore, these studies show that certain members of the FGF family have a very specialized biological role resulting in a highly specific phenotype (i.e. Angora mutant of FGF5
/
mice), while the loss of other FGFs can be compensated for by related members of the FGF family resulting in no obvious phenotypes (no obvious defect in FGF1
/
mice). Table 3 summarizes the phenotypes caused by targeted disruption of FGFR1–FGFR4 [32,39,74–80]. Targeted disruption of the FGFR1 causes embryonic lethality at E9.5–E12 as a result of defects in cell migration through the primitive streak. An identical phenotype was obtained by targeted disruption of the FGFR1c isoform, whereas no obvious phenotype was observed in FGFR1b
/
mice. It has been shown that FGFR2
/
mice die at E10.5 due to defects in the placenta. However, selective disruption of the FGFR2b isoform causes severe impairment in the development of the lung, limbs and other tissues resulting in lethality

Signaling via FGFRs is mediated via direct recruitment of signaling proteins that bind to tyrosine auto-phosphorylation sites on the activated receptor and via closely linked docking proteins that become tyrosine phosphorylated in response to FGF-stimulation and form a complex with additional complement of signaling proteins. The cytoplasmic domain of FGFR contains in addition to the catalytic PTK core, several regulatory sequences. The juxtamembrane domain of FGFRs is considerably longer than that of other receptor tyrosine kinases. This region contains a highly conserved sequence that serves as a binding site for the phosphotyrosine binding (PTB) domains of the two members of the FRS2 family of docking proteins FRS2a and FRS2b [27,28]. While the PTB domain of FRS2a or FRS2b binds to FGFR1 constitutively, independent of ligand stimulation and tyrosine phosphorylation, the same PTB domains bind to the juxtamembrane domain of NGF-receptor (TrkA) in a phosphorylation dependent manner to a canonical PTB domain-binding site (NPXpY motif). The tyrosine kinase domain of FGFRs is split like that of platelet-derived growth factor (PDGF) receptor or stem cell growth factor receptor (SCFR designated c-Kit), but the kinase insert region is much shorter in FGFRs than that of PDGFR and cKit. We have demonstrated that autophosphorylation on Tyr766 in the carboxy terminal tail of FGFR1 creates a specific-binding site for the SH2 domain of phospholipase Cg (PLCg) [29]. Mutational analysis of Y766 has shown

Table 2 Targeted disruption of FGF genes Gene

Survival

Phenotype

Reference

FGF1 FGF2 FGF3 FGF4 FGF5 FGF6 FGF7 FGF8 FGF9 FGF10 FGF14 FGF15 FGF17 FGF18 FGF23

Viable Viable Viable Lethal E5.5 Viable Viable Viable Lethal E8.5 Lethal Po Lethal Po Viable Viable Viable Lethal P1 Viable

No obvious phenotype Neuronal, skeletal and skin phenotypes Inner ear, tail outgrowth Inner cell mass proliferation Long hair, ‘‘Angora’’ phenotype Muscle regeneration Hair follicle and kidney deficiency Many phenotypes including gastrulation, brain, heart and craniofacial development Lung, xy sex reversal Many phenotypes including limbs, lungs, kidneys and others Neurological phenotype-ataxia and a paroxysmal hyperkinetic movement disorder No defect in inner ear development; poor survival rate Midline cerebral development Delayed ossification and increased chondrocyte proliferation; decreased alveolar spaces in the lung Hyperphosphatemia, hypoglycemia, reduced bone density and infertility

[56] [57] [58] [59] [60] [61] [62] [63] [64,65] [66,67] [68] [69] [53] [70–72] [73]

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

143

Table 3 Targeted mutagenesis of fibroblast growth factor receptor (FGFR) isotypes: loss of function Receptor/isoform

Survival

Phenotype

Reference

Fgfr1 Fgfr1b Fgfr1c Fgfr2 Fgfr2b Fgfr2c Fgfr3 Fgfr4

Lethal, Viable Lethal, Lethal, Lethal, Viable Viable Viable

Defective cell migration through primitive streak; posterior axis defect No obvious phenotype Defective cell migration through primitive streak; posterior axis defect Defect in placenta and limb bud formation Agenesis of lungs, anterior pituitary, thyroid, teeth and limbs Delayed ossification, proportionate dwarfism, synostosis of skull base (chondrocranium) Bone over growth; inner ear defect No obvious phenotype; growth retardation and lung defects in FGFR3 null background

[74,75] [32] [32] [76] [77] [78] [39,79] [80]

E9.5–E12.5 E9.5 E10.5 P0

that the phosphorylation of this tyrosine residue is essential for complex formation with and tyrosine phosphorylation of PLCg [30], resulting in PLCg activation, stimulation of phosphatidylinositol (PI) hydrolysis and the generation of the two second messengers, diacylglycerol and Ins(1,4,5)P3. Membrane recruitment of PLCg is aided by binding of the Pleckstrin homology (PH) domain of PLCg to PtdIns(3,4,5) P3 molecules that are generated in response to PI-3 kinase stimulation [31]. A mutant FGFR1 in which Y766 is replaced by phenylalanine is unable to activate PI hydrolysis and Ca2+ release in response to FGFstimulation suggesting that PI hydrolysis is dispensible for FGF-induced mitogenic stimulation of cultured cells. However, analysis of ‘‘knock-in’’ mice with mutated Y766 have shown that this tyrosine is required for a negative regulatory signal during anteroposterior patterning of mouse embryos [32]. Fig. 3 depicts the various signaling pathways activated by FGFRs.

Fig. 3. A wiring diagram depicting signaling pathways downstream of FGFR including the various signaling pathways that are dependent upon tyrosine phosphorylation of FRS2a following FGF-stimulation (green arrows). The negative signals that are mediated by or impinge upon FRS2a are marked by red arrows. Also shown, heterologous control of FGFsignaling by growth factors (i.e. insulin, EGF) or G-protein coupled agonists (i.e. LPA, carbachol) that stimulate threonine phosphorylation of FRS2a.

5.1. FRS2a functions as a major mediator of signaling via FGFR FGF-stimulation leads to tyrosine phosphorylation of the docking protein FRS2a and FRS2b, followed by recruitment of multiple Grb2/Sos complexes resulting in activation of the Ras/MAP kinase signaling pathway [33]. Tyrosine phosphorylated FRS2a functions as a site for coordinated assembly of a multiprotein complex that includes the docking protein Gab1 and the effector proteins that are recruited by this docking protein [34]. FRS2 proteins contain myristyl anchors and PTB domains in their N-termini and a large region with multiple tyrosine phosphorylation sites at their C-termini [33]. FRS2a contains four binding sites for the adaptor protein Grb2 and two binding sites for the protein tyrosine phosphatase Shp2. FGF-stimulation leads to tyrosine phosphorylation of Shp2 resulting in complex formation with additional Grb2 molecules. Grb2/Sos complexes are thus recruited directly and indirectly via Shp2 upon tyrosine phosphorylation of FRS2a in response to FGF-stimulation. The central role played by FRS2a in signaling via FGFRs was revealed by exploring FGFR signaling in fibroblasts isolated from FRS2a
/
embryos [34]. We have demonstrated that targeted disruption of the FRS2a gene causes severe impairment in mouse development resulting in embryonal lethality at E7–7.5. This result is consistent with earlier studies demonstrating that FGFR signaling plays critical roles at different stages of embryonic development (reviewed in [2,3]). As FRS2b is expressed exclusively in the nervous system of the embryo past E10–10.5, the second member of the family is unable to compensate for the loss of FRS2a earlier than E10 resulting in embryonic lethality at E7–7.5. Experiments with embryonic fibroblasts from FRS2a
/
mice demonstrate that FRS2a plays a critical role in FGFinduced MAP kinase stimulation, PI-3 kinase stimulation, chemotactic response and cell proliferation. We have also used fibroblasts isolated from FRS2a
/
embryos to demonstrate that FGF-induced tyrosine phosphorylation of the docking protein Gab1 depends on tyrosine phosphorylation of FRS2a. Gab1 binds constitutively to the Cterminal SH3 domain of Grb2 and its assembly in complex with Grb2/FRS2a enables tyrosine phosphorylation of

144

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

Gab1, which is followed by recruitment of a complement of SH2 domain containing signaling proteins including PI-3 kinase. FGF-induced recruitment of PI-3 kinase by Gab1 results in activation of the Akt dependent anti-apoptotic pathway. In addition to the central role played by FRS2a in recruitment and activation of the MAP kinase and PI-3 kinase, it has been shown that FRS2a plays a role in the recruitment of negative regulators [35]. Grb2 bound to tyrosine phosphorylated FRS2a forms a ternary complex with Cbl through the binding of the SH3 domains of Grb2 to a proline rich region in Cbl. Grb2-mediated recruitment of Cbl results in ubiquitination of FGFR and FRS2a. Cbl is a multidomain protein that posses an intrinsic ubiquitin ligase activity and also functions as a platform for recruitment of a variety of signaling proteins [35]. This experiment demonstrates that FRS2a is responsible for the assembly of both positive (i.e. Sos, PI-3 K) and negative (i.e. Cbl), signaling proteins to mediate a balanced FGF signal translation. In addition to enhancement of tyrosine phosphorylation, FGF-stimulation induces MAP kinase-dependent phosphorylation of FRS2a on at least eight threonine residues resulting in a large shift in its electrophoretic mobility [36]. Threonine phosphorylation of FRS2a is accompanied by reduced tyrosine phosphorylation of FRS2a, decreased recruitment of Grb2 and attenuation of the MAP kinase response. A similar FRS2a threonine phosphorylation is induced by PDGF, EGF or insulin stimulation, growth factors or hormones that do not induce tyrosine phosphorylation of FRS2a and do not stimulate the biological responses of FGFs. Prevention of FRS2a threonine phosphorylation by site directed mutagenesis or by treatment of the cells with the MEK inhibitor (PD 0980089) leads to constitutive tyrosine phosphorylation of FRS2a in unstimulated cells. Expression of an FRS2a mutant deficient in MAPK phosphorylation sites (the eight threonines have been replaced by valines, FRS2a-8V), induces anchorage independent cell growth and colony formation in soft agar; two hallmarks of cell transformation. In addition, FGF-induced tyrosine phosphorylation of FRS2a, MAP kinase stimulation and cell migration are strongly enhanced in FRS2a
/
cells expressing FRS2a8V mutant deficient in MAPK phosphorylation sites [36]. These experiments show that the same molecule that is responsible for the recruitment of positive regulators of signaling via FGFR can utilize a key element of the same pathway for a negative feedback mechanism resulting in signal attenuation and fine-tuning of its own activity [36].

6. Genetic alterations in FGFR genes in human disease 6.1. Skeletal disorders Several human skeletal dysplasias have been linked to specific point mutations in three members of the FGFR

family. It has been shown that point mutations in FGFR1, FGFR2 or FGFR3 are responsible for severe impairment in cranial, digital and skeletal development (reviewed in [37,38]). The most common craniosynostosis syndrome (premature fusion of cranial sutures) and skeletal dysplasia (dwarfism), have been linked to point mutations in FGFR1, 2 and 3. The mutations in FGFR1 that are responsible for Pfeifer syndrome and mutations in FGFR2 that are responsible for Pfiefer, Crouzon, Jackson–Weiss and Apert syndromes were summarized in Table 4 along with available animal models. Furthermore, point mutations in FGFR3 were linked to achondroplasia (ACH), hypochondroplasia (HCH), thanatophoric dysplasia type I and type II (TDI and TDII) (Table 4). The mutations responsible for HCH and TDII, are located in the catalytic PTK domain of FGFR. These are gain of function mutations that enhance the PTK activity in a ligand independent manner. The remaining gain-of-function mutations are confined to transmembrane or extracellular domains of FGFR. The most common form of human dwarfism is caused by a gain of function mutation in the transmembrane domain of FGFR3. Biochemical analyses confirmed that the ACH mutations increase both protein kinase activity and stability of the FGFR3 mutant protein. These results are consistent with the phenotype of FGFR3
/
mice. It was demonstrated that FGFR3 deficiency causes increased bone length due to chondrocyte hypertrophy [39]. Mutations in the extracellular domain of FGFRs cluster in three regions of the extracellular ligand-binding domain, in the linker connecting D2–D3, in D3 and in the region connecting D3 with the transmembrane domain. The large variety of gain of function mutations detected in these severe skeletal disorders activate the mutant FGFRs by either promoting FGFR dimerization or by altering ligand-receptor specificity. Many of the Crouzon, Pfiefer or Jackson–Weiss syndromes are caused by mutations in one of the two conserved cysteines in D3 of FGFR2, an amino acid residue that is normally linked intramolecularly to a second cysteine in the D3 of FGFR. The first group are mutations that substitute an amino acid with a cysteine residue or substitute a cysteine with another amino acid. Both types of mutations create an unpaired cysteine in the extracellular domain, which will form an intermolecular disulfide bridge, resulting in receptor dimerization and activation. The structure of the FGF/FGFR complexes suggested that many mutations in D3, although not directly involving cysteine residues, could destabilize the structure of D3 in such a way that certain cysteines that normally participate in the formation of intramolecular disulfide bridges will form instead intermolecular disulfide bridges with a cysteine residue in a neighboring receptor, again resulting in FGFR dimerization and activation. The second class of gain of function mutations that occur in the two highly conserved residues in the linker connecting D2 and D3 (Ser-252 and Pro-253), of FGFR2 are responsible for all known cases of Apert syndrome [37,38]. Both the structural information and

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

145

Table 4 Genetic alterations of FGFRs in human skeletal disorders Disease

Description

Gene

Activating mutation

Mouse models for human mutaion/ref

Crouzon syndrome

Synostosis of coronal sutures, midface hypoplacia, ocular proptosis

FGFR2

Cys342 Tyr; [81]

Jackson–Weiss syndrome Beare-Stevenson cutis gyrata

Craniosynostosis with foot abnormalities Cloverleaf skull, over growth of skin with furrowed palms and soles, prominent umbilical stump Severe and symmetric fusion of hands and feet, craniosynostosis Craniosynostosis, broad thumbs and toes

FGFR2 FGFR2; FGFR3

Multiple; about 39 different mutations were reported, e.g. Cys278Phe, Cys342Tyr, Ser347Cys Ala344Gly; Cys342Ser or Arg Ser372Cys; Tyr375Cys; Pro250Arg

FGFR2

Ser252Trp; Pro253Arg

Ser252Trp; [82]

FGFR1; FGFR2

Pro252Arg Multiple; 36 mutations reported Pro250Arg

Pro252Arg; [83]

Apert syndrome Pfeiffer syndrome Muenke syndrome Saethre-Chotzen-like syndrome Achondroplasia SADDAN (severe achondroplasia with developmental delay and acanthosis nigricans) Thanatophoric dysplasia type I Thanatophoric dysplasia type II Hypochondroplasia

Unilateral or bilateral coronal synostosis; thimble like middle phalanges (hands) Craniofacial and limb defects Short statue, midface deficiency

FGFR3

Short limbs, developmental delay, acanthosis nigricans

FGFR3

Curved short femurs; lethal

FGFR3

Stright femurs with cloverleaf skull; lethal Short limbs but less severe than achondroplasia, large head circumference

FGFR3 FGFR3

FGFR2; FGFR3 FGFR3

ValVal269-70del Pro250Arg Gly346Glu; Gly375Cys; Gly380Arg Lys650Met

Multiple; 9 different mutations reported Lys650Glu Multiple; 9 different mutations reported

Gly380Arg; [84] Gly375Cys; [85] Lys650Met; [86]

Ser371Cys; [87] Lys650Glu; [88]

For complete listing of mutations, see [89] and [90].

ligand-binding experiments indicate that these mutations cause the mesenchymal splice form of FGFR2 (FGFR2c) to bind and to be activated by the mesenchymally expressed ligands FGF7 or FGF10 and the epithelial splice form of FGFR2 (FGFR2b) to be activated by FGF2, FGF6 and FGF9 [40,41].

6.2. Kallmann syndrome Kallmann syndrome (KAL1) is a developmental disease in which the GnRH-synthesizing neurons fail to migrate from olfactory epithelium causing hypogonadism and ansonia (absent or underdeveloped olfactory bulb) [42].

Table 5 Genetic alterations of FGFRs in human cancers Cancer

Gene alteration

Reference

8P11 myeloproliferative syndrome (EMS)

Translocation and fusion of FGFR1 with ZNF-198 (also called FIM or RAMP) t(8;13); fusion of FOP with FGFR1 t(6;8); fusion of FGFR1 with CEP 110 t(8;9); fusion of FGFR1 with endogenous human retroviral sequence t(8;19); and fusion of FGFR1 with BCR t(8;22) Over expression of FGFR1 Abnormal exprssion of FGFR1 and FGFR4 Class switch of FGFR2 from IIIb isoform to IIIc isoform Abnormal expression of FGFR1c in prostate epithelial cells Elevated expression of FGFR1 in white matter and down regulation of FGFR2 in malignant astrocytomas FGFR2 splice site mutation (940-2A ! G) and Ser267Pro mutation Frequent FGFR3 mutations: Arg248Cys; Ser249Cys; Gly372Cys; Lys652Glu Over expression of FGFR3 Low frequency of FGFR3 mutation: Ser249Cys Aberrant splicing and activation of cryptic splice sequences in FGFR3 Tanslocation and fusion of FGFR3 with ETV6 t(4;12) Activating mutations of FGFR3 (Lys650Glu; Lys650Met) associated with chromosomal translocation t(4:14) (p16.3;q32.3) FGFR4 polymorphism: Gly388Arg

[91–94]; [95]; [96]; [97]; [47,50]

Breast cancer Pancreatic adenocarcinoma Prostate cancer Astrocytoma Gastric cancer Transitional cell carcinoma of bladder Thyroid carcinoma Cervical carcinoma Colorectal cancer Peripheral T cell lymphoma Multiple myeloma Head and neck squamous cell carcinoma

[98] [99,100] [101] [102] [103] [51,104–106] [107] [51,105,108] [103,109] [110] [111] [112]

146

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

The frequently associated symptoms are hearing loss, tooth agenesis and cleft lip/palate [42]. The gene responsible for this syndrome is KAL1, which encodes ansonin-1, modular glycoprotein that is present in various extracellular matrices [43,44]. Recently, it has been found that somatic mutations of FGFR1 causes autosomal dominant form of Kallmann syndrome (KAL2) [45]. So far 15 different mutations of FGFR1 were reported for KAL2 syndrome [42]. The majority of the mutations are heterozygos leading to the impairment of FGFR function. These findings are consistent with the observation that mice deficient in Fgfr1 expression in telencephalon fail to develop the olfactory bulb, a characteristic feature of KAL2 syndrome [46]. 6.3. Cancer Constitutive activation of tyrosine kinases as fusion proteins with other genes due to chromosomal translocations plays important role in the development of many hematological malignancies especially in myeloproliferative syndromes (MPS). Translocation and fusion of FGFR1 with other genes cause a specific type of MPS syndrome called ‘8p11 myeloproliferative syndrome’ (EMS) which is characterized by myeloid hyperplasia, eosinophilia and lymphoblastic lymphoma [47,48]. So far, five different fusion partners for FGFR1 have been identified (Table 5). These fusion proteins induce constitutive tyrosine kinase activation of FGFR1 by oligomerization. For example, in the case of ZNF-198-FGFR1 fusion protein, the tyrosine kinase activation requires a novel proline rich oligomerization domain in ZNF-198. Although the majority of FGF signals are transduced via the docking protein FRS2, these fusion kinases seem to utilize pathways that are independent of FRS2. For instance, the ZNF-198-FGFR1 and Bcr-FGFR1 fusion kinases lack the juxtamembrane domain, which is required for FRS2 binding [49]. However, these kinases utilize Tyr 766 to recruit PLC-g which is absolutely required for the transforming activity of the fusion kinases [49]. The Bcr-FGFR1 tyrosine kinase recruits, in addition to PLC-g, Grb2 through phospho Tyr177 present on Bcr. This additional pathway produced a distinct form of myeloproliferative disorder different from EMS but closely related to chronic myeloid leukemia (CML) [50] suggesting that the outcome of the disease may be dependent on the signaling pathways that are activated by the chimeric FGFR protein [49]. Similar translocation and fusion of FGFR3 kinases are associated with multiple myeloma (MM) and peripheral T cell lymphoma. Table 5 summarizes the various types of cancers that are associated with altered expression of FGFRs. Germline mutations that are associated with achondroplasia and thantophoric dysplasia are associated with 35% of cases of bladder cancer and 25% of cases of cervical carcinoma [51]. Although, FGFR3 is associated with a large number of human cancers, much has to be done to elucidate the downstream signaling pathways responsible for FGFR3 kinase induced cell transformation.

Acknowledgement The laboratory of J. Schlessinger is supported by NIH grant RO1-AR051448 and by funds from the Ludwig Institute for Cancer Research.

References [1] Gospodarowicz D. Localisation of a fibroblast growth factor and its effect alone and with hydrocortisone on 3T3 cell growth. Nature 1974;249:123–7. [2] Ornitz DM, Itoh N. Fibroblast growth factors. Genome Biol 2001;2:3005 [Reviews]. [3] Givol D, Eswarakumar VP, Lonai P. Molecular and cellular biology of FGF signaling. In: Epstein CJ, Erickson RP, Wynshaw-Boris A, editors. Inborn errors of development – the molecular basis of clinical disorders of morphogenesis. Oxford: Oxford University Press; 2003. p. 367–79. [4] Lee PL, Johnson DE, Cousens LS, Fried VA, Williams LT. Purification and complementary DNA cloning of a receptor for basic fibroblast growth factor. Science 1989;245:57–60. [5] Givol D, Yayon A. Complexity of FGF receptors: genetic basis for structural diversity and functional specificity. FASEB J 1992;6:3362– 9. [6] Jaye M, Schlessinger J, Dionne CA. Fibroblast growth factor receptor tyrosine kinases: molecular analysis and signal transduction. Biochim Biophys Acta 1992;1135:185–99. [7] Yayon A, Klagsbrun M, Esko JD, Leder P, Ornitz DM. Cell surface, heparin-like molecules are required for binding of basic fibroblast growth factor to its high affinity receptor. Cell 1991;64:841–8. [8] Rapraeger AC, Krufka A, Olwin BB. Requirement of heparan sulfate for bFGF-mediated fibroblast growth and myoblast differentiation. Science 1991;252:1705–8. [9] Ornitz DM, Yayon A, Flanagan JG, Svahn CM, Levi E, Leder P. Heparin is required for cell-free binding of basic fibroblast growth factor to a soluble receptor and for mitogenesis in whole cells. Mol Cell Biol 1992;12:240–7. [10] Spivak-Kroizman T, Lemmon MA, Dikic I, Ladbury JE, Pinchasi D, Huang J, et al. Heparin-induced oligomerization of FGF molecules is responsible for FGF receptor dimerization, activation, and cell proliferation. Cell 1994;79:1015–24. [11] Lin X, Buff EM, Perrimon N, Michelson AM. Heparan sulfate proteoglycans are essential for FGF receptor signaling during Drosophila embryonic development. Development 1999;126: 3715–23. [12] Hunter T. Signaling—2000 and beyond. Cell 2000;100:113–27. [13] Schlessinger J. Cell signaling by receptor tyrosine kinases. Cell 2000;103:211–25. [14] Schlessinger J, Plotnikov AN, Ibrahimi OA, Eliseenkova AV, Yeh BK, Yayon A, et al. Crystal structure of a ternary FGF–FGFR– heparin complex reveals a dual role for heparin in FGFR binding and dimerization. Mol Cell 2000;6:743–50. [15] Miki T, Bottaro DP, Fleming TP, Smith CL, Burgess WH, Chan AM, et al. Determination of ligand-binding specificity by alternative splicing: two distinct growth factor receptors encoded by a single gene. Proc Natl Acad Sci USA 1992;89:246–50. [16] Yayon A, Zimmer Y, Shen GH, Avivi A, Yarden Y, Givol D. A confined variable region confers ligand specificity on fibroblast growth factor receptors: implications for the origin of the immunoglobulin fold. EMBO J 1992;11:1885–90. [17] Orr-Urtreger A, Bedford MT, Burakova T, Arman E, Zimmer Y, Yayon A, et al. Developmental localization of the splicing alternatives of fibroblast growth factor receptor-2 (FGFR2). Dev Biol 1993;158:475–86.

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149 [18] Plotnikov AN, Schlessinger J, Hubbard SR, Mohammadi M. Structural basis for FGF receptor dimerization and activation. Cell 1999;98:641–50. [19] Plotnikov AN, Hubbard SR, Schlessinger J, Mohammadi M. Crystal structures of two FGF–FGFR complexes reveal the determinants of ligand–receptor specificity. Cell 2000;101:413–24. [20] Wang F, Kan M, Yan G, Xu J, McKeehan WL. Alternately spliced NH2-terminal immunoglobulin-like Loop I in the ectodomain of the fibroblast growth factor (FGF) receptor 1 lowers affinity for both heparin and FGF-1. J Biol Chem 1995;270:10231–5. [21] Olsen SK, Ibrahimi OA, Raucci A, Zhang F, Eliseenkova AV, Yayon A, et al. Insights into the molecular basis for fibroblast growth factor receptor autoinhibition and ligand-binding promiscuity. Proc Natl Acad Sci USA 2004;101:935–40. [22] Schlessinger J. Signal transduction. Autoinhibition control. Science 2003;300:750–2. [23] Mohammadi M, Schlessinger J, Hubbard SR. Structure of the FGF receptor tyrosine kinase domain reveals a novel autoinhibitory mechanism. Cell 1996;86:577–87. [24] Mohammadi M, McMahon G, Sun L, Tang C, Hirth P, Yeh BK, et al. Structures of the tyrosine kinase domain of fibroblast growth factor receptor in complex with inhibitors. Science 1997;276:955–60. [25] Mohammadi M, Froum S, Hamby JM, Schroeder MC, Panek RL, Lu GH, et al. Crystal structure of an angiogenesis inhibitor bound to the FGF receptor tyrosine kinase domain. EMBO J 1998;17:5896–904. [26] Noble ME, Endicott JA, Johnson LN. Protein kinase inhibitors: insights into drug design from structure. Science 2004;303:1800–5. [27] Ong SH, Guy GR, Hadari YR, Laks S, Gotoh N, Schlessinger J, et al. FRS2 proteins recruit intracellular signaling pathways by binding to diverse targets on fibroblast growth factor and nerve growth factor receptors. Mol Cell Biol 2000;20:979–89. [28] Dhalluin C, Yan K, Plotnikova O, Lee KW, Zeng L, Kuti M, et al. Structural basis of SNT PTB domain interactions with distinct neurotrophic receptors. Mol Cell 2000;6:921–9. [29] Mohammadi M, Honegger AM, Rotin D, Fischer R, Bellot F, Li W, et al. A tyrosine-phosphorylated carboxy-terminal peptide of the fibroblast growth factor receptor (Flg) is a binding site for the SH2 domain of phospholipase C-gamma 1. Mol Cell Biol 1991;11:5068–78. [30] Mohammadi M, Dionne CA, Li W, Li N, Spivak T, Honegger AM, et al. Point mutation in FGF receptor eliminates phosphatidylinositol hydrolysis without affecting mitogenesis. Nature 1992;358:681–4. [31] Falasca M, Logan SK, Lehto VP, Baccante G, Lemmon MA, Schlessinger J. Activation of phospholipase C gamma by PI 3-kinaseinduced PH domain-mediated membrane targeting. EMBO J 1998;17:414–22. [32] Partanen J, Schwartz L, Rossant J. Opposite phenotypes of hypomorphic and Y766 phosphorylation site mutations reveal a function for Fgfr1 in anteroposterior patterning of mouse embryos. Genes Dev 1998;12:2332–44. [33] Kouhara H, Hadari YR, Spivak-Kroizman T, Schilling J, Bar-Sagi D, Lax I, et al. A lipid-anchored Grb2-binding protein that links FGFreceptor activation to the Ras/MAPK signaling pathway. Cell 1997;89:693–702. [34] Hadari YR, Gotoh N, Kouhara H, Lax I, Schlessinger J. Critical role for the docking-protein FRS2 alpha in FGF receptor-mediated signal transduction pathways. Proc Natl Acad Sci USA 2001;98:8578–83. [35] Wong A, Lamothe B, Lee A, Schlessinger J, Lax I, Li A. FRS2 alpha attenuates FGF receptor signaling by Grb2-mediated recruitment of the ubiquitin ligase Cbl. Proc Natl Acad Sci USA 2002;99:6684–9. [36] Lax I, Wong A, Lamothe B, Lee A, Frost A, Hawes J, et al. The docking protein FRS2alpha controls a MAP kinase-mediated negative feedback mechanism for signaling by FGF receptors. Mol Cell 2002;10:709–19. [37] Webster MK, Donoghue DJ. FGFR activation in skeletal disorders: too much of a good thing. Trends Genet 1997;13:178–82. [38] Wilkie AO. Craniosynostosis: genes and mechanisms. Hum Mol Genet 1997;6:1647–56.

147

[39] Deng C, Wynshaw-Boris A, Zhou F, Kuo A, Leder P. Fibroblast growth factor receptor 3 is a negative regulator of bone growth. Cell 1996;84:911–21. [40] Yu K, Herr AB, Waksman G, Ornitz DM. Loss of fibroblast growth factor receptor 2 ligand-binding specificity in Apert syndrome. Proc Natl Acad Sci USA 2000;97:14536–41. [41] Ibrahimi OA, Eliseenkova AV, Plotnikov AN, Yu K, Ornitz DM, Mohammadi M. Structural basis for fibroblast growth factor receptor 2 activation in Apert syndrome. Proc Natl Acad Sci USA 2001;98:7182–7. [42] Dode C, Hardelin JP. Kallmann syndrome: fibroblast growth factor signaling insufficiency? J Mol Med 2004;82:725–34. [43] Franco B, Guioli S, Pragliola A, Incerti B, Bardoni B, Tonlorenzi R, et al. A gene deleted in Kallmann’s syndrome shares homology with neural cell adhesion and axonal path-finding molecules. Nature 1991;353:529–36. [44] Legouis R, Hardelin JP, Levilliers J, Claverie JM, Compain S, Wunderle V, et al. The candidate gene for the X-linked Kallmann syndrome encodes a protein related to adhesion molecules. Cell 1991;67:423–35. [45] Dode C, Levilliers J, Dupont JM, De Paepe A, Le Du N, SoussiYanicostas N, et al. Loss-of-function mutations in FGFR1 cause autosomal dominant Kallmann syndrome. Nat Genet 2003;33: 463–5. [46] Hebert JM, Lin M, Partanen J, Rossant J, McConnell SK. FGF signaling through FGFR1 is required for olfactory bulb morphogenesis. Development 2003;130:1101–11. [47] Fioretos T, Panagopoulos I, Lassen C, Swedin A, Billstrom R, Isaksson M, et al. Fusion of the BCR and the fibroblast growth factor receptor-1 (FGFR1) genes as a result of t(8;22)(p11;q11) in a myeloproliferative disorder: the first fusion gene involving BCR but not ABL. Genes Chromosomes Cancer 2001;32:302–10. [48] Macdonald D, Aguiar RC, Mason PJ, Goldman JM, Cross NC. A new myeloproliferative disorder associated with chromosomal translocations involving 8p11: a review. Leukemia 1995;9:1628–30. [49] Roumiantsev S, Krause DS, Neumann CA, Dimitri CA, Asiedu F, Cross NC, et al. Distinct stem cell myeloproliferative/T lymphoma syndromes induced by ZNF198–FGFR1 and BCR–FGFR1 fusion genes from 8p11 translocations. Cancer Cell 2004;5: 287–98. [50] Demiroglu A, Steer EJ, Heath C, Taylor K, Bentley M, Allen SL, et al. The t(8;22) in chronic myeloid leukemia fuses BCR to FGFR1: transforming activity and specific inhibition of FGFR1 fusion proteins. Blood 2001;98:3778–83. [51] Cappellen D, De Oliveira C, Ricol D, de Medina S, Bourdin J, SastreGarau X, et al. Frequent activating mutations of FGFR3 in human bladder and cervix carcinomas. Nat Genet 1999;23:18–20. [52] Ornitz DM, Xu J, Colvin JS, McEwen DG, MacArthur CA, Coulier F, et al. Receptor specificity of the fibroblast growth factor family. J Biol Chem 1996;271:15292–7. [53] Xu J, Liu Z, Ornitz DM. Temporal and spatial gradients of Fgf8 and Fgf17 regulate proliferation and differentiation of midline cerebellar structures. Development 2000;127:1833–43. [54] Umemori H, Linhoff MW, Ornitz DM, Sanes JR. FGF22 and its close relatives are presynaptic organizing molecules in the mammalian brain. Cell 2004;118:257–70. [55] Yamashita T, Konishi M, Miyake A, Inui K, Itoh N. Fibroblast growth factor (FGF)-23 inhibits renal phosphate reabsorption by activation of the mitogen-activated protein kinase pathway. J Biol Chem 2002;277:28265–70. [56] Miller DL, Ortega S, Bashayan O, Basch R, Basilico C. Compensation by fibroblast growth factor 1 (FGF1) does not account for the mild phenotypic defects observed in FGF2 null mice. Mol Cell Biol 2000;20:2260–8. [57] Dono R, Texido G, Dussel R, Ehmke H, Zeller R. Impaired cerebral cortex development and blood pressure regulation in FGF-2-deficient mice. EMBO J 1998;17:4213–25.

148

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149

[58] Mansour SL, Goddard JM, Capecchi MR. Mice homozygous for a targeted disruption of the proto-oncogene int-2 have developmental defects in the tail and inner ear. Development 1993;117: 13–28. [59] Feldman B, Poueymirou W, Papaioannou VE, DeChiara TM, Goldfarb M. Requirement of FGF-4 for postimplantation mouse development. Science 1995;267:246–9. [60] Hebert JM, Rosenquist T, Gotz J, Martin GR. FGF5 as a regulator of the hair growth cycle: evidence from targeted and spontaneous mutations. Cell 1994;78:1017–25. [61] Floss T, Arnold HH, Braun T. A role for FGF-6 in skeletal muscle regeneration. Genes Dev 1997;11:2040–51. [62] Guo L, Degenstein L, Fuchs E. Keratinocyte growth factor is required for hair development but not for wound healing. Genes Dev 1996;10:165–75. [63] Meyers EN, Lewandoski M, Martin GR. An Fgf8 mutant allelic series generated by Cre- and Flp-mediated recombination. Nat Genet 1998;18:136–41. [64] Colvin JS, White AC, Pratt SJ, Ornitz DM. Lung hypoplasia and neonatal death in Fgf9-null mice identify this gene as an essential regulator of lung mesenchyme. Development 2001;128: 2095–106. [65] Colvin JS, Green RP, Schmahl J, Capel B, Ornitz DM. Male-tofemale sex reversal in mice lacking fibroblast growth factor 9. Cell 2001;104:875–89. [66] Min H, Danilenko DM, Scully SA, Bolon B, Ring BD, Tarpley JE, et al. Fgf-10 is required for both limb and lung development and exhibits striking functional similarity to Drosophila branchless. Genes Dev 1998;12:3156–61. [67] Sekine K, Ohuchi H, Fujiwara M, Yamasaki M, Yoshizawa T, Sato T, et al. Fgf10 is essential for limb and lung formation. Nat Genet 1999;21:138–41. [68] Wang Q, Bardgett ME, Wong M, Wozniak DF, Lou J, McNeil BD, et al. Ataxia and paroxysmal dyskinesia in mice lacking axonally transported FGF14. Neuron 2002;35:25–38. [69] Wright TJ, Ladher R, McWhirter J, Murre C, Schoenwolf GC, Mansour SL. Mouse FGF15 is the ortholog of human and chick FGF19, but is not uniquely required for otic induction. Dev Biol 2004;269:264–75. [70] Liu Z, Xu J, Colvin JS, Ornitz DM. Coordination of chondrogenesis and osteogenesis by fibroblast growth factor 18. Genes Dev 2002;16:859–69. [71] Ohbayashi N, Shibayama M, Kurotaki Y, Imanishi M, Fujimori T, Itoh N, et al. FGF18 is required for normal cell proliferation and differentiation during osteogenesis and chondrogenesis. Genes Dev 2002;16:870–9. [72] Usui H, Shibayama M, Ohbayashi N, Konishi M, Takada S, Itoh N. Fgf18 is required for embryonic lung alveolar development. Biochem Biophys Res Commun 2004;322:887–92. [73] Shimada T, Kakitani M, Yamazaki Y, Hasegawa H, Takeuchi Y, Fujita T, et al. Targeted ablation of Fgf23 demonstrates an essential physiological role of FGF23 in phosphate and vitamin D metabolism. J Clin Invest 2004;113:561–8. [74] Deng CX, Wynshaw-Boris A, Shen MM, Daugherty C, Ornitz DM, Leder P. Murine FGFR-1 is required for early postimplantation growth and axial organization. Genes Dev 1994;8:3045–57. [75] Yamaguchi TP, Harpal K, Henkemeyer M, Rossant J. fgfr-1 is required for embryonic growth and mesodermal patterning during mouse gastrulation. Genes Dev 1994;8:3032–44. [76] Xu X, Weinstein M, Li C, Naski M, Cohen RI, Ornitz DM, et al. Fibroblast growth factor receptor 2 (FGFR2)-mediated reciprocal regulation loop between FGF8 and FGF10 is essential for limb induction. Development 1998;125:753–65. [77] De Moerlooze L, Spencer-Dene B, Revest J, Hajihosseini M, Rosewell I, Dickson C. An important role for the IIIb isoform of fibroblast growth factor receptor 2 (FGFR2) in mesenchymal–epithelial signalling during mouse organogenesis. Development 2000;127:483–92.

[78] Eswarakumar VP, Monsonego-Ornan E, Pines M, Antonopoulou I, Morriss-Kay GM, Lonai P. The IIIc alternative of Fgfr2 is a positive regulator of bone formation. Development 2002;129: 3783–93. [79] Colvin JS, Bohne BA, Harding GW, McEwen DG, Ornitz DM. Skeletal overgrowth and deafness in mice lacking fibroblast growth factor receptor 3. Nat Genet 1996;12:390–7. [80] Weinstein M, Xu X, Ohyama K, Deng CX. FGFR-3 and FGFR-4 function cooperatively to direct alveogenesis in the murine lung. Development 1998;125:3615–23. [81] Eswarakumar VP, Horowitz MC, Locklin R, Morriss-Kay GM, Lonai P. A gain-of-function mutation of Fgfr2c demonstrates the roles of this receptor variant in osteogenesis. Proc Natl Acad Sci USA 2004;101:12555–60. [82] Chen L, Li D, Li C, Engel A, Deng CX. A Ser250Trp substitution in mouse fibroblast growth factor receptor 2 (Fgfr2) results in craniosynostosis. Bone 2003;33:169–78. [83] Zhou YX, Xu X, Chen L, Li C, Brodie SG, Deng CX. A Pro250Arg substitution in mouse Fgfr1 causes increased expression of Cbfa1 and premature fusion of calvarial sutures. Hum Mol Genet 2000;9:2001– 8. [84] Wang Y, Spatz MK, Kannan K, Hayk H, Avivi A, Gorivodsky M, et al. A mouse model for achondroplasia produced by targeting fibroblast growth factor receptor 3. Proc Natl Acad Sci USA 1999;96:4455–60. [85] Chen L, Adar R, Yang X, Monsonego EO, Li C, Hauschka PV, et al. Gly369Cys mutation in mouse FGFR3 causes achondroplasia by affecting both chondrogenesis and osteogenesis. J Clin Invest 1999;104:1517–25. [86] Iwata T, Li CL, Deng CX, Francomano CA. Highly activated Fgfr3 with the K644M mutation causes prolonged survival in severe dwarf mice. Hum Mol Genet 2001;10:1255–64. [87] Chen L, Li C, Qiao W, Xu X, Deng C. A Ser(365) ! Cys mutation of fibroblast growth factor receptor 3 in mouse downregulates Ihh/ PTHrP signals and causes severe achondroplasia. Hum Mol Genet 2001;10:457–65. [88] Iwata T, Chen L, Li C, Ovchinnikov DA, Behringer RR, Francomano CA, et al. A neonatal lethal mutation in FGFR3 uncouples proliferation and differentiation of growth plate chondrocytes in embryos. Hum Mol Genet 2000;9:1603–13. [89] Passos-Bueno MR, Wilcox WR, Jabs EW, Sertie AL, Alonso LG, Kitoh H. Clinical spectrum of fibroblast growth factor receptor mutations. Hum Mutat 1999;14:115–25. [90] Cohen Jr MM. FGFs/FGFRs and associated disorders. In: Epstein CJ, Erickson RP, Wynshaw-Boris A, editors. Inborn errors of development—the molecular basis of clinical disorders of morphogenesis. Oxford: Oxford University Press; 2003. p. 380–400. [91] Popovici C, Adelaide J, Ollendorff V, Chaffanet M, Guasch G, Jacrot M, et al. Fibroblast growth factor receptor 1 is fused to FIM in stemcell myeloproliferative disorder with t(8;13). Proc Natl Acad Sci USA 1998;95:5712–7. [92] Reiter A, Sohal J, Kulkarni S, Chase A, Macdonald DH, Aguiar RC, et al. Consistent fusion of ZNF198 to the fibroblast growth factor receptor-1 in the t(8;13)(p11;q12) myeloproliferative syndrome. Blood 1998;92:1735–42. [93] Smedley D, Hamoudi R, Clark J, Warren W, Abdul-Rauf M, Somers G, et al. The t(8;13)(p11;q11-12) rearrangement associated with an atypical myeloproliferative disorder fuses the fibroblast growth factor receptor 1 gene to a novel gene RAMP. Hum Mol Genet 1998;7:637–42. [94] Xiao S, Nalabolu SR, Aster JC, Ma J, Abruzzo L, Jaffe ES, et al. FGFR1 is fused with a novel zinc-finger gene, ZNF198, in the t(8;13) leukaemia/lymphoma syndrome. Nat Genet 1998;18:84–7. [95] Popovici C, Zhang B, Gregoire MJ, Jonveaux P, Lafage-Pochitaloff M, Birnbaum D, et al. The t(6;8)(q27;p11) translocation in a stem cell myeloproliferative disorder fuses a novel gene, FOP, to fibroblast growth factor receptor 1. Blood 1999;93:1381–9.

V.P. Eswarakumar et al. / Cytokine & Growth Factor Reviews 16 (2005) 139–149 [96] Guasch G, Mack GJ, Popovici C, Dastugue N, Birnbaum D, Rattner JB, et al. FGFR1 is fused to the centrosome-associated protein CEP110 in the 8p12 stem cell myeloproliferative disorder with t(8;9)(p12;q33). Blood 2000;95:1788–96. [97] Guasch G, Popovici C, Mugneret F, Chaffanet M, Pontarotti P, Birnbaum D, et al. Endogenous retroviral sequence is fused to FGFR1 kinase in the 8p12 stem-cell myeloproliferative disorder with t(8;19)(p12;q13.3). Blood 2003;101:286–8. [98] Yoshimura N, Sano H, Hashiramoto A, Yamada R, Nakajima H, Kondo M, et al. The expression and localization of fibroblast growth factor-1 (FGF-1) and FGF receptor-1 (FGFR-1) in human breast cancer. Clin Immunol Immunopathol 1998;89:28–34. [99] Kobrin MS, Yamanaka Y, Friess H, Lopez ME, Korc M. Aberrant expression of type I fibroblast growth factor receptor in human pancreatic adenocarcinomas. Cancer Res 1993;53: 4741–4. [100] Shah RN, Ibbitt JC, Alitalo K, Hurst HC. FGFR4 overexpression in pancreatic cancer is mediated by an intronic enhancer activated by HNF1alpha. Oncogene 2002;21:8251–61. [101] Kwabi-Addo B, Ropiquet F, Giri D, Ittmann M. Alternative splicing of fibroblast growth factor receptors in human prostate cancer. Prostate 2001;46:163–72. [102] Yamaguchi F, Saya H, Bruner JM, Morrison RS. Differential expression of two fibroblast growth factor-receptor genes is associated with malignant progression in human astrocytomas. Proc Natl Acad Sci USA 1994;91:484–8. [103] Jang JH, Shin KH, Park JG. Mutations in fibroblast growth factor receptor 2 and fibroblast growth factor receptor 3 genes associated with human gastric and colorectal cancers. Cancer Res 2001;61:3541–3. [104] Kimura T, Suzuki H, Ohashi T, Asano K, Kiyota H, Eto Y. The incidence of thanatophoric dysplasia mutations in FGFR3 gene is

[105]

[106]

[107]

[108]

[109]

[110]

[111]

[112]

149

higher in low-grade or superficial bladder carcinomas. Cancer 2001;92:2555–61. Sibley K, Stern P, Knowles MA. Frequency of fibroblast growth factor receptor 3 mutations in sporadic tumours. Oncogene 2001;20:4416–8. van Rhijn BW, Lurkin I, Radvanyi F, Kirkels WJ, van der Kwast TH, Zwarthoff EC. The fibroblast growth factor receptor 3 (FGFR3) mutation is a strong indicator of superficial bladder cancer with low recurrence rate. Cancer Res 2001;61:1265–8. Onose H, Emoto N, Sugihara H, Shimizu K, Wakabayashi I. Overexpression of fibroblast growth factor receptor 3 in a human thyroid carcinoma cell line results in overgrowth of the confluent cultures. Eur J Endocrinol 1999;140:169–73. Wu R, Connolly D, Ngelangel C, Bosch FX, Munoz N, Cho KR. Somatic mutations of fibroblast growth factor receptor 3 (FGFR3) are uncommon in carcinomas of the uterine cervix. Oncogene 2000;19:5543–6. Jang JH, Shin KH, Park YJ, Lee RJ, McKeehan WL, Park JG. Novel transcripts of fibroblast growth factor receptor 3 reveal aberrant splicing and activation of cryptic splice sequences in colorectal cancer. Cancer Res 2000;60:4049–52. Yagasaki F, Wakao D, Yokoyama Y, Uchida Y, Murohashi I, Kayano H, et al. Fusion of ETV6 to fibroblast growth factor receptor 3 in peripheral T-cell lymphoma with a t(4;12)(p16;p13) chromosomal translocation. Cancer Res 2001;61:8371–4. Chesi M, Nardini E, Brents LA, Schrock E, Ried T, Kuehl WM, et al. Frequent translocation t(4;14)(p16.3;q32.3) in multiple myeloma is associated with increased expression and activating mutations of fibroblast growth factor receptor 3. Nat Genet 1997;16:260–4. Streit S, Bange J, Fichtner A, Ihrler S, Issing W, Ullrich A. Involvement of the FGFR4 Arg388 allele in head and neck squamous cell carcinoma. Int J Cancer 2004;111:213–7.