Добірка наукової літератури з теми "Fgf-receptors biosynthesis"

Farnesyl pyrophosphate synthase (FPPS) catalyses the formation of a key cellular intermediate in isoprenoid metabolic pathways. Here we describe a novel role for this enzyme in fibroblast growth factor (FGF)-mediated signalling. We demonstrate the binding of FPPS to FGF receptors (FGFRs) using the yeast two-hybrid assay, pull-down assays and co-immunoprecipitation. The interaction between FPPS and FGFR is regulated by the cellular metabolic state and by treatment with FGF-2. Overexpression of FPPS inhibits FGF-2-induced cell proliferation, accompanied by a failure of the FGF-2-mediated induction of cyclins D1 and E. Overexpression of FPPS in fibroblasts also promotes increased farnesylation of Ras, and temporally extends FGF-2-stimulated activation of the Ras/ERK (extracellular-signal-regulated kinase) cascade. These data suggest that, in addition to its role in isoprenoid biosynthesis, FPPS may function as a modulator of the cellular response to FGF treatment.

Several peptide growth factors, including members of the fibroblast growth factor (FGF) superfamily, are potential inducers of mesoderm in vertebrates. Receptor binding of basic FGF (FGF-2) is promoted by cell surface or extracellular matrix proteoglycans. The substantial biosynthesis of proteoglycans by embryonic cells (called embryoglycans) and their potential role as ligands for growth factor receptors led us to examine the role of embryoglycans that carry the developmentally regulated oligosaccharide epitope TEC 1, in the binding of FGF-2 to cultured rabbit inner cell masses (ICMs). Culture of isolated ICMs in the presence of FGF-2 gave rise to well delimited colonies with migrating cells at the periphery. In these cells, TEC 1 staining shifts from a punctate pattern over the entire membrane, to an apical, finely granular distribution with some internalization. This shift occurs after 96 hours in culture. Here we show that: (1) migrating cells are mesoderm-like in phenotype; (2) antibodies against TEC 1 blocked FGF-2 mediated differentiation in vitro; (3) antibodies against TEC 1 selectively blocked binding of FGF-2 to ectodermal receptors and, vice versa, the binding of TEC 1-specific antibodies to ectodermal cells can be competed by excess FGF-2; (4) the same switch in TEC 1 staining patterns was observed in vivo, between the day 7 and the day 9 rabbit embryo. These data suggest the involvement of defined species of embryonic cell surface epitopes in the regulation of FGF-2 receptor binding. Moreover, this proposed binding activity is temporally restricted to ectodermal cells and disappears early during differentiation. Thus, the apical TEC 1 redistribution can be considered as the earliest indicator of mesoderm formation.

Abstract The klotho gene functions as an aging-suppressor gene that extends life span when overexpressed and accelerates aging-like phenotypes when disrupted in mice. The klotho gene encodes a single-pass transmembrane protein that binds to multiple fibroblast growth factor (FGF) receptors and functions as a co-receptor for FGF23, a bone-derived hormone that suppresses phosphate reabsorption and vitamin D biosynthesis in the kidney. In addition, the extracellular domain of Klotho protein is shed and secreted, potentially functioning as a humoral factor. The secreted Klotho protein can regulate multiple growth factor signaling pathways, including insulin/IGF-1 and Wnt, and the activity of multiple ion channels. Klotho protein also protects cells and tissues from oxidative stress, yet the precise mechanism underlying these activities remains to be determined. Thus, understanding of Klotho protein function is expected to provide new insights into the molecular basis for aging, phosphate/vitamin D metabolism, cancer and stem cell biology.

The Drosophila sugarless and sulfateless genes encode enzymes required for the biosynthesis of heparan sulfate glycosaminoglycans. Biochemical studies have shown that heparan sulfate glycosaminoglycans are involved in signaling by fibroblast growth factor receptors, but evidence for such a requirement in an intact organism has not been available. We now demonstrate that sugarless and sulfateless mutant embryos have phenotypes similar to those lacking the functions of two Drosophila fibroblast growth factor receptors, Heartless and Breathless. Moreover, both Heartless- and Breathless-dependent MAPK activation is significantly reduced in embryos which fail to synthesize heparan sulfate glycosaminoglycans. Consistent with an involvement of Sulfateless and Sugarless in fibroblast growth factor receptor signaling, a constitutively activated form of Heartless partially rescues sugarless and sulfateless mutants, and dosage-sensitive interactions occur between heartless and the heparan sulfate glycosaminoglycan biosynthetic enzyme genes. We also find that overexpression of Branchless, the Breathless ligand, can partially overcome the requirement of Sugarless and Sulfateless for Breathless activity. These results provide the first genetic evidence that heparan sulfate glycosaminoglycans are essential for fibroblast growth factor receptor signaling in a well defined developmental context, and support a model in which heparan sulfate glycosaminoglycans facilitate fibroblast growth factor ligand and/or ligand-receptor oligomerization.

I recettori per i fattori di crescita dei fibroblasti (FGFR) presentano attività tirosino chinasica (TK) e vengono processati attraverso differenti fasi di maturazione/glicosilazione che avvengono nei compartimenti cellulari della via di secrezione. In questo studio abbiamo determinato due nuovi eventi che caratterizzano la biosintesi dell’FGFR3. Innanzitutto, dimostriamo che una limitata frazione del recettore selvaggio immaturo, ricco in mannosi, è sottoposta a dimerizzazione transeunte nelle fasi precoci della via secretoria. Diversamente, le forme glicosilate mature, proseguono verso la superficie cellulare come monomeri. L’efficienza di dimerizzazione del recettore è aumentata dall’attività chinasica costitutiva associata alla mutazione K650E del FGFR3, ritenuto nel reticolo endoplasmico (ER) e responsabile della displasia tanatoforica di tipo II (TDII). Dimostriamo, infatti che l’abrogazione di tale attività chinasica riporta l’efficienza di dimerizzazione ai livelli basali osservati per il recettore selvaggio. Dimostriamo, inoltre, che il mutante K650E dimerizza e trans-fosforila l’FGFR3 selvaggio nella sua forma immatura supportando l’ipotesi che la dimerizzazione tra il recettore mutato e quello selvaggio comporti il sequestro di quest’ ultimo nel ER. Tale evento potrebbe spiegare il carattere genetico “dominante” dell’allele K650E nella TDII. Per mezzo della tecnica di “complementazione dei frammenti proteici basata sulla ricostituzione della fluorescenza” confermiamo la dimerizzazione del FGFR3 in vivo. Mostriamo inoltre che l’FGFR3 è una proteina cargo per la lectina mannosio-specifica ERGIC-53. Nell’insieme questi dati evidenziano che la dimerizzazione transeunte è un evento nella biosintesi dell’FGFR3 ed ipotizziamo possa fare parte di un meccanismo di “controllo qualità” di preassemblaggio del recettore, dal momento che l’attivazione funzionale del FGFR3 avviene sotto forma dimerica ed è conseguente all’interazione con il ligando specifico FGF, sulla superficie cellulare. Mostriamo inoltre, per la prima volta la presenza di complessi ERGIC-53/FGFR3 suggerendo un ruolo per ERGIC53 nel trasporto di FGFR3 dal ER al Golgi. Osserviamo infine che anche l’FGFR2, un altro membro della famiglia degli FGFR, presenta un livello ‘basale’ di dimerizzazione delle forme immature ricche in mannosio, suggerendo che tale dimerizzazione intracellulare possa essere un meccanismo comune a tutti i membri della famiglia degli FGFRs.
The tyrosine kinase Fibroblast Growth Factor Receptors (FGFRs) are processed though different stages of maturation/glycosylation occurring in the various compartments of the secretory pathway. In this study, we have determined a novel event that characterizes the biosynthesis of FGFR3. We show that a limited fraction of the immature mannose-rich wild-type receptor undergoes transient dimerization in the early phase of the secretory pathway. The mature FGFR3 glycomers however, pursues the cell surface as monomers. The constitutive kinase activity associated to the K650E FGFR3 mutant that is retained in the endoplasmic reticulum (ER), increases dimerization efficiency and accordingly, abrogation of the kinase activity restores the “basal” dimerization level observed for the wild-type FGFR3. Furthermore, the K650E mutant dimerizes and trans-phosphorylates the immature wild-type FGFR3 supporting the hypothesis that dimerization of mutant and wild-type FGFR3 causes the retention of the latter in the ER. This may explain the genetic dominance of the K650E allele in the TDII disease. Fluorescence-based protein fragment complementation assay confirms FGFR3 dimerization in vivo and indicates FGFR3 as a cargo of the mannose-specific lectin ERGIC-53. Altogether these data highlight transient dimerization as a step in the FGFR3 biosynthesis and we hypothesize that this event could be part of a ‘pre-assembly quality control’ of the receptor, since the receptor function on the cell surface is exerted through the homo-dimerization upon the interaction with the specific FGF ligand. Furthermore, we show for the first time the presence of ERGIC53/FGFR3 complexes, suggesting a role for ERGIC53 in the transport of the FGFR3 from the ER to the Golgi. Finally, we observed that FGFR2, another member of the FGFR family, also shows a ‘basal’ level of dimerization of the immature mannose-rich isoforms, suggesting that intracellular dimerization could be a mechanism shared by all the members of the FGFRs family.