Fibroblast growth factors (FGFs) control organ morphogenesis during development as well as tissue homeostasis and repair in the adult organism. Despite their importance, many mechanisms that regulate FGF function are still poorly understood. Interestingly, the thermodynamic stability of 22 mammalian FGFs varies widely, with some FGFs remaining stable at body temperature for more than 24 h, while others lose their activity within minutes. How thermodynamic stability contributes to the function of FGFs during development remains unknown. Here we show that FGF10, an important limb and lung morphogen, exists as an intrinsically unstable protein that is prone to unfolding and is rapidly inactivated at 37 °C. Using rationally driven directed mutagenesis, we have developed several highly stable (STAB) FGF10 variants with a melting temperature of over 19 °C more than that of wildtype FGF10. In cellular assays in vitro, the FGF10-STABs did not differ from wildtype FGF10 in terms of binding to FGF receptors, activation of downstream FGF receptor signaling in cells, and induction of gene expression. In mouse embryonal lung explants, FGF10-STABs, but not wildtype FGF10, suppressed branching, resulting in increased alveolarization and expansion of epithelial tissue. Similarly, FGF10-STAB1, but not FGF10 wildtype, inhibited the growth of mouse embryonic tibias and markedly altered limb morphogenesis when implanted into chicken limb buds, collectively demonstrating that thermal instability should be considered an important regulator of FGF function that prevents ectopic signaling. Furthermore, we show enhanced differentiation of human iPSC-derived lung organoids and improved regeneration in ex vivo lung injury models mediated by FGF10-STABs, suggesting an application in cell therapy.

• Klíčová slova
• Development, FGF10, Fibroblast growth factor, Lung, Morphogen, Stability,
• MeSH
• fibroblastový růstový faktor 10 * metabolismus   genetika   chemie   MeSH
• lidé MeSH
• myši MeSH
• plíce metabolismus   embryologie   MeSH
• receptory fibroblastových růstových faktorů metabolismus   MeSH
• signální transdukce * MeSH
• stabilita proteinů MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• Fgf10 protein, mouse MeSH Prohlížeč
• fibroblastový růstový faktor 10 * MeSH
• receptory fibroblastových růstových faktorů MeSH

Achondroplasia is the most common form of human dwarfism caused by mutations in the FGFR3 receptor tyrosine kinase. Current therapy begins at 2 years of age and improves longitudinal growth but does not address the cranial malformations including midface hypoplasia and foramen magnum stenosis, which lead to significant otolaryngeal and neurologic compromise. A recent clinical trial found partial restoration of cranial defects with therapy starting at 3 months of age, but results are still inconclusive. The benefits of achondroplasia therapy are therefore controversial, increasing skepticism among the medical community and patients. We used a mouse model of achondroplasia to test treatment protocols aligned with human studies. Early postnatal treatment (from day 1) was compared with late postnatal treatment (from day 4, equivalent to ~5 months in humans). Animals were treated with the FGFR3 inhibitor infigratinib and the effect on skeleton was thoroughly examined. We show that premature fusion of the skull base synchondroses occurs immediately after birth and leads to defective cranial development and foramen magnum stenosis in the mouse model to achondroplasia. This phenotype appears significantly restored by early infigratinib administration when compared with late treatment, which provides weak to no rescue. In contrast, the long bone growth is similarly improved by both early and late protocols. We provide clear evidence that immediate postnatal therapy is critical for normalization of skeletal growth in both the cranial base and long bones and the prevention of sequelae associated with achondroplasia. We also describe the limitations of early postnatal therapy, providing a paradigm-shifting argument for the development of prenatal therapy for achondroplasia.

The article provides clear evidence that achondroplasia should be treated immediately after birth, not only to increase height (appendicular growth), but more importantly to prevent defective cranial skeletogenesis and associated severe neurological complications. Although later treatment promotes growth of the long bones (achondroplasia patients grow taller), the defective head skeleton that forms before and/or early after birth cannot be restored if therapy is not started immediately after birth. We also describe the limitations of postnatal treatment and make a strong case for the development of prenatal therapy for achondroplasia, which appears necessary for a comprehensive treatment of this condition.

• Klíčová slova
• Fgfr3, achondroplasia, fibroblast growth factor, infigratinib, postnatal, treatment,
• MeSH
• achondroplazie * patologie   farmakoterapie   MeSH
• lebka patologie   účinky léků   MeSH
• lidé MeSH
• modely nemocí na zvířatech * MeSH
• myši MeSH
• receptor fibroblastových růstových faktorů, typ 3 * genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• receptor fibroblastových růstových faktorů, typ 3 * MeSH

The FGF system is the most complex of all receptor tyrosine kinase signaling networks with 18 FGF ligands and four FGFRs that deliver morphogenic signals to pattern most embryonic structures. Even when a single FGFR is expressed in the tissue, different FGFs can trigger dramatically different biological responses via this receptor. Here we show both quantitative and qualitative differences in the signaling of one of the FGF receptors, FGFR1c, in response to different FGFs. We provide an overview of the recent discovery that FGFs engage in biased signaling via FGFR1c. We discuss the concept of ligand bias, which represents qualitative differences in signaling as it is a measure of differential ligand preferences for different downstream responses. We show how FGF ligand bias manifests in functional data in cultured chondrocyte cells. We argue that FGF-ligand bias contributes substantially to FGF-driven developmental processes, along with known differences in FGF expression levels, FGF-FGFR binding coefficients and differences in FGF stability in vivo.

• Klíčová slova
• Bias, FGF, FGFR, Signaling,
• MeSH
• chondrocyty metabolismus   MeSH
• fibroblastové růstové faktory * metabolismus   MeSH
• lidé MeSH
• ligandy MeSH
• receptor fibroblastových růstových faktorů, typ 1 * metabolismus   MeSH
• receptory fibroblastových růstových faktorů * metabolismus   MeSH
• signální transdukce * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• fibroblastové růstové faktory * MeSH
• ligandy MeSH
• receptor fibroblastových růstových faktorů, typ 1 * MeSH
• receptory fibroblastových růstových faktorů * MeSH

The differential signaling of multiple FGF ligands through a single fibroblast growth factor (FGF) receptor (FGFR) plays an important role in embryonic development. Here, we use quantitative biophysical tools to uncover the mechanism behind differences in FGFR1c signaling in response to FGF4, FGF8, and FGF9, a process which is relevant for limb bud outgrowth. We find that FGF8 preferentially induces FRS2 phosphorylation and extracellular matrix loss, while FGF4 and FGF9 preferentially induce FGFR1c phosphorylation and cell growth arrest. Thus, we demonstrate that FGF8 is a biased FGFR1c ligand, as compared to FGF4 and FGF9. Förster resonance energy transfer experiments reveal a correlation between biased signaling and the conformation of the FGFR1c transmembrane domain dimer. Our findings expand the mechanistic understanding of FGF signaling during development and bring the poorly understood concept of receptor tyrosine kinase ligand bias into the spotlight.

• Klíčová slova
• FGFR, biased signaling, molecular biophysics, none, signal transduction, structural biology,
• MeSH
• fibroblastové růstové faktory * MeSH
• fosforylace MeSH
• lidé MeSH
• ligandy MeSH
• receptor fibroblastových růstových faktorů, typ 1 genetika   MeSH
• signální transdukce * MeSH
• těhotenství MeSH
• zkreslení výsledků (epidemiologie) MeSH
• Check Tag
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• FGFR1 protein, human MeSH Prohlížeč
• fibroblastové růstové faktory * MeSH
• ligandy MeSH
• receptor fibroblastových růstových faktorů, typ 1 MeSH

Successful specification of the two mouse blastocyst inner cell mass (ICM) lineages (the primitive endoderm (PrE) and epiblast) is a prerequisite for continued development and requires active fibroblast growth factor 4 (FGF4) signaling. Previously, we identified a role for p38 mitogen-activated protein kinases (p38-MAPKs) during PrE differentiation, but the underlying mechanisms have remained unresolved. Here, we report an early blastocyst window of p38-MAPK activity that is required to regulate ribosome-related gene expression, rRNA precursor processing, polysome formation and protein translation. We show that p38-MAPK inhibition-induced PrE phenotypes can be partially rescued by activating the translational regulator mTOR. However, similar PrE phenotypes associated with extracellular signal-regulated kinase (ERK) pathway inhibition targeting active FGF4 signaling are not affected by mTOR activation. These data indicate a specific role for p38-MAPKs in providing a permissive translational environment during mouse blastocyst PrE differentiation that is distinct from classically reported FGF4-based mechanisms.

• MeSH
• blastocysta fyziologie   MeSH
• buněčná diferenciace MeSH
• buněčný rodokmen MeSH
• DNA vazebné proteiny fyziologie   MeSH
• embryonální vývoj MeSH
• endoderm cytologie   MeSH
• mitogenem aktivované proteinkinasy p38 antagonisté a inhibitory   fyziologie   MeSH
• myši MeSH
• proteiny vázající RNA fyziologie   MeSH
• proteosyntéza * MeSH
• TOR serin-threoninkinasy fyziologie   MeSH
• transkripční faktory fyziologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA vazebné proteiny MeSH
• mitogenem aktivované proteinkinasy p38 MeSH
• mTOR protein, mouse MeSH Prohlížeč
• Mybbp1a protein, mouse MeSH Prohlížeč
• proteiny vázající RNA MeSH
• TOR serin-threoninkinasy MeSH
• transkripční faktory MeSH

Fibroblast growth factors (FGFs) deliver extracellular signals that govern many developmental and regenerative processes, but the mechanisms regulating FGF signaling remain incompletely understood. Here, we explored the relationship between intrinsic stability of FGF proteins and their biological activity for all 18 members of the FGF family. We report that FGF1, FGF3, FGF4, FGF6, FGF8, FGF9, FGF10, FGF16, FGF17, FGF18, FGF20, and FGF22 exist as unstable proteins, which are rapidly degraded in cell cultivation media. Biological activity of FGF1, FGF3, FGF4, FGF6, FGF8, FGF10, FGF16, FGF17, and FGF20 is limited by their instability, manifesting as failure to activate FGF receptor signal transduction over long periods of time, and influence specific cell behavior in vitro and in vivo. Stabilization via exogenous heparin binding, introduction of stabilizing mutations or lowering the cell cultivation temperature rescues signaling of unstable FGFs. Thus, the intrinsic ligand instability is an important elementary level of regulation in the FGF signaling system.

• MeSH
• chondrosarkom genetika   metabolismus   patologie   MeSH
• cirkulární dichroismus MeSH
• fibroblastové růstové faktory chemie   klasifikace   genetika   metabolismus   MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• mutace genetika   MeSH
• mutantní proteiny chemie   metabolismus   MeSH
• nádorové buňky kultivované MeSH
• nádory kostí genetika   metabolismus   patologie   MeSH
• nádory prsu genetika   metabolismus   patologie   MeSH
• proliferace buněk * MeSH
• signální transdukce * MeSH
• stabilita proteinů MeSH
• teplota MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fibroblastové růstové faktory MeSH
• mutantní proteiny MeSH

In 1994, the field of bone biology was significantly advanced by the discovery that activating mutations in the fibroblast growth factor receptor 3 (FGFR3) receptor tyrosine kinase (TK) account for the common genetic form of dwarfism in humans, achondroplasia (ACH). Other conditions soon followed, with the list of human disorders caused by FGFR3 mutations now reaching at least 10. An array of vastly different diagnoses is caused by similar mutations in FGFR3, including syndromes affecting skeletal development (hypochondroplasia [HCH], ACH, thanatophoric dysplasia [TD]), skin (epidermal nevi, seborrhaeic keratosis, acanthosis nigricans), and cancer (multiple myeloma [MM], prostate and bladder carcinoma, seminoma). Despite many years of research, several aspects of FGFR3 function in disease remain obscure or controversial. As FGFR3-related skeletal dysplasias are caused by growth attenuation of the cartilage, chondrocytes appear to be unique in their response to FGFR3 activation. However, the reasons why FGFR3 inhibits chondrocyte growth while causing excessive cellular proliferation in cancer are not clear. Likewise, the full spectrum of molecular events by which FGFR3 mediates its signaling is just beginning to emerge. This article describes the challenging journey to unravel the mechanisms of FGFR3 function in skeletal dysplasias, the extraordinary cellular manifestations of FGFR3 signaling in chondrocytes, and finally, the progress toward therapy for ACH and cancer.

• MeSH
• chondrocyty metabolismus   patologie   MeSH
• chrupavka abnormality   metabolismus   MeSH
• fibroblastové růstové faktory genetika   metabolismus   MeSH
• fosfatidylinositol-3-kinasy genetika   metabolismus   MeSH
• kosti a kostní tkáň abnormality   metabolismus   MeSH
• kůže metabolismus   patologie   MeSH
• letální geny MeSH
• lidé MeSH
• MAP kinasový signální systém genetika   MeSH
• mezibuněčná komunikace MeSH
• mutace MeSH
• nádory kůže genetika   metabolismus   patologie   MeSH
• natriuretický peptid typu C genetika   metabolismus   MeSH
• osteochondrodysplazie genetika   metabolismus   patologie   MeSH
• proliferace buněk MeSH
• receptor fibroblastových růstových faktorů, typ 3 * genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• signální transdukce MeSH
• transkripční faktor STAT1 genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• FGFR3 protein, human MeSH Prohlížeč
• fibroblastové růstové faktory MeSH
• fosfatidylinositol-3-kinasy MeSH
• natriuretický peptid typu C MeSH
• receptor fibroblastových růstových faktorů, typ 3 * MeSH
• STAT1 protein, human MeSH Prohlížeč
• transkripční faktor STAT1 MeSH

Oncogenic activation of the RAS-ERK MAP kinase signaling pathway can lead to uncontrolled proliferation but can also result in apoptosis or premature cellular senescence, both regarded as natural protective barriers to cell immortalization and transformation. In FGFR3-related skeletal dyplasias, oncogenic mutations in the FGFR3 receptor tyrosine kinase cause profound inhibition of cartilage growth resulting in severe dwarfism, although many of the precise mechanisms of FGFR3 action remain unclear. Mutated FGFR3 induces constitutive activation of the ERK pathway in chondrocytes and, remarkably, can also cause both increased proliferation and apoptosis in growing cartilage, depending on the gestational age. Here, we demonstrate that FGFR3 signaling is also capable of inducing premature senescence in chondrocytes, manifested as reversible, ERK-dependent growth arrest accompanied by alteration of cellular shape, loss of the extracellular matrix, upregulation of senescence markers (alpha-GLUCOSIDASE, FIBRONECTIN, CAVEOLIN 1, LAMIN A, SM22alpha and TIMP 1), and induction of senescence-associated beta-GALACTOSIDASE activity. Our data support a model whereby FGFR3 signaling inhibits cartilage growth via exploiting cellular responses originally designed to eliminate cells harboring activated oncogenes.

• MeSH
• apoptóza MeSH
• chondrocyty enzymologie   patologie   MeSH
• extracelulární matrix metabolismus   MeSH
• extracelulárním signálem regulované MAP kinasy metabolismus   MeSH
• fenotyp MeSH
• krysa rodu Rattus MeSH
• onkogeny genetika   MeSH
• proliferace buněk MeSH
• receptor fibroblastových růstových faktorů, typ 3 metabolismus   MeSH
• signální transdukce * MeSH
• stárnutí buněk * MeSH
• tvar buňky MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• extracelulárním signálem regulované MAP kinasy MeSH
• receptor fibroblastových růstových faktorů, typ 3 MeSH

Activation of fibroblast growth factor receptor 3 (FGFR3) leads to attenuation of cartilage growth. The members of the STAT family of transcription factors are believed to participate in FGFR3 signaling in cartilage, however the molecular mechanism of this action is poorly understood. Here, we demonstrate that a chronic FGF stimulus leads to accumulation of STAT1, 3, 5 and 6, evident in both in vitro chondrocyte model and murine limb explant cultures. Despite the accumulation, both endogenous and cytokine-induced activation of STAT1 and STAT3 is impaired by FGF, as demonstrated by imaging of active STAT nuclear translocation and analyses of STAT activatory phosphorylation and transcriptional activation. Further, we demonstrate that FGF induces expression of CIS, SOCS1 and SOCS3 inhibitors of gp130, a common receptor for the IL6-family of cytokines. Since cytokine-gp130 signaling represents an important positive regulator of cartilage, its inhibition may contribute to the growth-inhibitory effect of FGFR3 in cartilage.

• MeSH
• chondrocyty účinky léků   metabolismus   MeSH
• cytokinový receptor gp130 metabolismus   MeSH
• fibroblastový růstový faktor 2 farmakologie   MeSH
• interferon gama antagonisté a inhibitory   farmakologie   MeSH
• interleukin-6 antagonisté a inhibitory   farmakologie   MeSH
• myši MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• protein SOCS1 MeSH
• protein SOCS3 MeSH
• proteiny SOCS metabolismus   MeSH
• receptor fibroblastových růstových faktorů, typ 3 metabolismus   MeSH
• sekvence nukleotidů MeSH
• signální transdukce MeSH
• transkripční faktor STAT1 antagonisté a inhibitory   metabolismus   MeSH
• transkripční faktor STAT3 antagonisté a inhibitory   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• cytokine inducible SH2-containing protein MeSH Prohlížeč
• cytokinový receptor gp130 MeSH
• fibroblastový růstový faktor 2 MeSH
• interferon gama MeSH
• interleukin-6 MeSH
• protein SOCS1 MeSH
• protein SOCS3 MeSH
• proteiny SOCS MeSH
• receptor fibroblastových růstových faktorů, typ 3 MeSH
• Socs1 protein, mouse MeSH Prohlížeč
• Socs3 protein, mouse MeSH Prohlížeč
• transkripční faktor STAT1 MeSH
• transkripční faktor STAT3 MeSH

Activating mutations in FGFR3 tyrosine kinase cause several forms of human skeletal dysplasia. Although the mechanisms of FGFR3 action in cartilage are not completely understood, it is believed that the STAT1 transcription factor plays a central role in pathogenic FGFR3 signaling. Here, we analyzed STAT1 activation by the N540K, G380R, R248C, Y373C, K650M and K650E-FGFR3 mutants associated with skeletal dysplasias. In a cell-free kinase assay, only K650M and K650E-FGFR3 caused activatory STAT1(Y701) phosphorylation. Similarly, in RCS chondrocytes, HeLa, and 293T cellular environments, only K650M and K650E-FGFR3 caused strong STAT1 activation. Other FGFR3 mutants caused weak (HeLa) or no activation (293T and RCS). This contrasted with ERK MAP kinase activation, which was strongly induced by all six mutants and correlated with the inhibition of proliferation in RCS chondrocytes. Thus the ability to activate STAT1 appears restricted to the K650M and K650E-FGFR3 mutants, which however account for only a small minority of the FGFR3-related skeletal dysplasia cases. Other pathways such as ERK should therefore be considered as central to pathological FGFR3 signaling in cartilage.

• MeSH
• bezbuněčný systém metabolismus   MeSH
• biologické modely MeSH
• CHO buňky MeSH
• chrupavka metabolismus   MeSH
• Cricetulus MeSH
• extracelulárním signálem regulované MAP kinasy metabolismus   MeSH
• fosforylace MeSH
• HeLa buňky MeSH
• kosti a kostní tkáň patologie   MeSH
• křečci praví MeSH
• krysa rodu Rattus MeSH
• kultivované buňky MeSH
• lidé MeSH
• mutantní proteiny fyziologie   MeSH
• receptor fibroblastových růstových faktorů, typ 3 genetika   fyziologie   MeSH
• signální transdukce genetika   fyziologie   MeSH
• transkripční faktor STAT1 analýza   metabolismus   fyziologie   MeSH
• vývojové onemocnění kostí genetika   metabolismus   patologie   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• extracelulárním signálem regulované MAP kinasy MeSH
• mutantní proteiny MeSH
• receptor fibroblastových růstových faktorů, typ 3 MeSH
• transkripční faktor STAT1 MeSH