scholarly journals A designed fusion tag for soluble expression and selective separation of extracellular domains of fibroblast growth factor receptors

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Dae-Eun Cheong ◽  
Hye-Ji Choi ◽  
Su-Kyoung Yoo ◽  
Hun-Dong Lee ◽  
Geun-Joong Kim
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Expression Patterns ◽  
Growth Factor Receptors ◽  
Selective Separation ◽  
Soluble Expression ◽  
Terminal Region ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Extracellular Domains

AbstractFibroblast growth factor receptors (FGFRs) generate various transduction signals by interaction with fibroblast growth factors (FGFs) and are involved in various biological functions such as cell proliferation, migration, and differentiation. Malfunction of these proteins may lead to the development of various diseases, including cancer. Accordingly, FGFRs are considered an alternative therapeutic target for protein and/or gene therapy. However, the screening of antagonists or agonists of FGFRs is challenging due to their complex structural features associated with protein expression. Herein, we conducted the development of a protease-free cleavable tag (PFCT) for enhancing the solubility of difficult-to express protein by combining maltose-binding protein (MBP) and the C-terminal region of Npu intein. To validate the availability of the resulting tag for the functional production of extracellular domains of FGFRs (Ec_FGFRs), we performed fusion of PFCT with the N-terminus of Ec_FGFRs and analyzed the expression patterns. Almost all PFCT-Ec_FGFR fusion proteins were mainly detected in the soluble fraction except for Ec_FGFR4. Upon addition of the N-terminal region of Npu intein, approximately 85% of the PFCT-Ec_FGFRs was separated into PFCT and Ec_FGFR via intein-mediated cleavage. Additionally, the structural integrity of Ec_FGFR was confirmed by affinity purification using heparin column. Taken together, our study demonstrated that the PFCT could be used for soluble expression and selective separation of Ec_FGFRs.

Heterogeneity in the expression of fibroblast growth factor receptors during limb regeneration in newts (Notophthalmus viridescens)

Development ◽  
1993 ◽  
Vol 119 (2) ◽  
pp. 353-361 ◽  
Author(s):  
M.L. Poulin ◽  
K.M. Patrie ◽  
M.J. Botelho ◽  
R.A. Tassava ◽  
I.M. Chiu
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Expression Patterns ◽  
Limb Regeneration ◽  
Basal Layer ◽  
Growth Factor Receptors ◽  
Notophthalmus Viridescens ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Wound Epithelium

Two closely related fibroblast growth factor receptors, FGFR1 and FGFR2, have been cloned from a newt (Notophthalmus viridescens) limb blastema cDNA library. Sequence analysis revealed that we have isolated both the bek and KGFR variants of FGFR2. These two variants differ only in the second half of the last of their three Ig-like domains. The expression patterns of FGFR1 and FGFR2 during limb regeneration have been determined by in situ hybridization. During the preblastema stages of regeneration, FGFR2 expression is observed in the basal layer of the wound epithelium and in the cells of the periosteum. As regeneration progresses to the blastema stages, FGFR2 expression continues to be observed in the basal layer of the wound epithelium with additional hybridization seen in the blastema mesenchyme closely associated with the bisected bones. From the early bud to the mid-bud blastema stage, FGFR1 expression is observed throughout the blastema mesenchyme but, unlike FGFR2, is distinctly absent from the wound epithelium. In the differentiation stages of regeneration, the mesenchymal expression of FGFR2 becomes restricted to the cells of the condensing cartilage and later to the perichondrium. During these later stages of regeneration, the wound epithelium hybridization to the FGFR2 probe is no longer observed. The expression patterns of these receptors suggest that FGFR1 and FGFR2 have distinct roles in limb regeneration, despite their sharing a number of the FGF ligands. Further investigation regarding the potential sources of the FGF ligands will help establish the role that FGFs and FGFRs play in limb regeneration.

scholarly journals Targeting fibroblast growth factor receptors to combat aggressive ependymoma

2021 ◽  
Author(s):  
Daniela Lötsch ◽  
Dominik Kirchhofer ◽  
Bernhard Englinger ◽  
Li Jiang ◽  
Konstantin Okonechnikov ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Molecular Mechanisms ◽  
Radial Glia ◽  
Growth Factor Receptors ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Fibroblast Growth ◽  
Cell Models ◽  
Fibroblast Growth Factor Receptors

AbstractEpendymomas (EPN) are central nervous system tumors comprising both aggressive and more benign molecular subtypes. However, therapy of the high-risk subtypes posterior fossa group A (PF-A) and supratentorial RELA-fusion positive (ST-RELA) is limited to gross total resection and radiotherapy, as effective systemic treatment concepts are still lacking. We have recently described fibroblast growth factor receptors 1 and 3 (FGFR1/FGFR3) as oncogenic drivers of EPN. However, the underlying molecular mechanisms and their potential as therapeutic targets have not yet been investigated in detail. Making use of transcriptomic data across 467 EPN tissues, we found that FGFR1 and FGFR3 were both widely expressed across all molecular groups. FGFR3 mRNA levels were enriched in ST-RELA showing the highest expression among EPN as well as other brain tumors. We further identified high expression levels of fibroblast growth factor 1 and 2 (FGF1, FGF2) across all EPN subtypes while FGF9 was elevated in ST-EPN. Interrogation of our EPN single-cell RNA-sequencing data revealed that FGFR3 was further enriched in cycling and progenitor-like cell populations. Corroboratively, we found FGFR3 to be predominantly expressed in radial glia cells in both mouse embryonal and human brain datasets. Moreover, we detected alternative splicing of the FGFR1/3-IIIc variant, which is known to enhance ligand affinity and FGFR signaling. Dominant-negative interruption of FGFR1/3 activation in PF-A and ST-RELA cell models demonstrated inhibition of key oncogenic pathways leading to reduced cell growth and stem cell characteristics. To explore the feasibility of therapeutically targeting FGFR, we tested a panel of FGFR inhibitors in 12 patient-derived EPN cell models revealing sensitivity in the low-micromolar to nano-molar range. Finally, we gain the first clinical evidence for the activity of the FGFR inhibitor nintedanib in the treatment of a patient with recurrent ST-RELA. Together, these preclinical and clinical data suggest FGFR inhibition as a novel and feasible approach to combat aggressive EPN.

Semirational design of a potent, artificial agonist of fibroblast growth factor receptors

10.1038/70746 ◽  
1999 ◽  
Vol 17 (12) ◽  
pp. 1199-1204 ◽  
Author(s):  
Marcus D. Ballinger ◽  
Venkatakrishna Shyamala ◽  
Louise D. Forrest ◽  
Maja Deuter-Reinhard ◽  
Laura V. Doyle ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Growth Factor Receptors ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors

scholarly journals Role of Klotho in Hyperglycemia: Its Levels and Effects on Fibroblast Growth Factor Receptors, Glycolysis, and Glomerular Filtration

2021 ◽  
Vol 22 (15) ◽  
pp. 7867
Author(s):  
Marlena Typiak ◽  
Tomasz Kulesza ◽  
Patrycja Rachubik ◽  
Dorota Rogacka ◽  
Irena Audzeyenka ◽  
...  
Keyword(s):  
Growth Factor ◽  
Urinary Excretion ◽  
Glomerular Filtration ◽  
Fibroblast Growth Factor ◽  
Growth Factor Receptors ◽  
Renal Tissue ◽  
Proper Function ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Plasma Filtration

Hyperglycemic conditions (HG), at early stages of diabetic nephropathy (DN), cause a decrease in podocyte numbers and an aberration of their function as key cells for glomerular plasma filtration. Klotho protein was shown to overcome some negative effects of hyperglycemia. Klotho is also a coreceptor for fibroblast growth factor receptors (FGFRs), the signaling of which, together with a proper rate of glycolysis in podocytes, is needed for a proper function of the glomerular filtration barrier. Therefore, we measured levels of Klotho in renal tissue, serum, and urine shortly after DN induction. We investigated whether it influences levels of FGFRs, rates of glycolysis in podocytes, and albumin permeability. During hyperglycemia, the level of membrane-bound Klotho in renal tissue decreased, with an increase in the shedding of soluble Klotho, its higher presence in serum, and lower urinary excretion. The addition of Klotho increased FGFR levels, especially FGFR1/FGFR2, after their HG-induced decrease. Klotho also increased levels of glycolytic parameters of podocytes, and decreased podocytic and glomerular albumin permeability in HG. Thus, we found that the decrease in the urinary excretion of Klotho might be an early biomarker of DN and that Klotho administration may have several beneficial effects on renal function in DN.

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