scholarly journals Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3512 ◽  
Author(s):  
Adam Bajinting ◽  
Ho Leung Ng
Keyword(s):  
Tyrosine Kinases ◽  
Recombinant Expression ◽  
Expression System ◽  
Transmembrane Helix ◽  
Size Exclusion ◽  
Complex Signal ◽  
Tyrosine Kinase Domain ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
E Coli

Fibroblast growth factor receptors (FGFRs) are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs) as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

scholarly journals Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

2017 ◽  
Author(s):  
Adam Bajinting ◽  
Ho Leung Ng
Keyword(s):  
Tyrosine Kinases ◽  
Recombinant Expression ◽  
Expression System ◽  
Transmembrane Helix ◽  
Size Exclusion ◽  
Complex Signal ◽  
Tyrosine Kinase Domain ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
E Coli

ABSTRACTFibroblast growth factor receptors (FGFRs) are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs) as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

Cell transformation by fibroblast growth factors can be suppressed by truncated fibroblast growth factor receptors

1994 ◽  
Vol 14 (11) ◽  
pp. 7660-7669
Author(s):  
Y Li ◽  
C Basilico ◽  
A Mansukhani
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Tyrosine Kinases ◽  
Dominant Negative ◽  
Receptor Expression ◽  
Scatchard Analysis ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Truncated Receptor ◽  
Nih 3T3

Ligand-induced dimerization and transphosphorylation are thought to be important events by which receptor tyrosine kinases generate cellular signals. We have investigated the ability of signalling-defective, truncated fibroblast growth factor (FGF) receptors (FGFR-1 and FGFR-2) to block the FGF response in cells that express both types of endogenous FGF receptors. When these dominant negative receptors are expressed in NIH 3T3 cells transformed by the secreted FGF-4, the transformed properties of the cells can be reverted to various degrees, with better reversion phenotype correlating with higher levels of truncated receptor expression. Furthermore, truncated FGFR-2 is significantly more efficient at producing reversion than FGFR-1, indicating that FGF-4 preferentially utilizes the FGFR-2 signalling pathway. NIH 3T3 clones expressing these truncated receptors are more resistant to FGF-induced mitogenesis and also exhibit reduced tyrosine phosphorylation upon treatment with FGF. The block in FGF-signalling, however, can be overcome by the addition of excess growth factor. The truncated receptors have binding affinities that are four- to eightfold lower than those of wild-type receptors, as measured by Scatchard analysis. We also observed a partial specificity in the responses of truncated-receptor-expressing clones to FGF-2 or FGF-4. Our results suggest that the block to signal transduction produced by kinase-negative FGF receptors is achieved through a combination of dominant negative effects and competition for growth factor binding with functional receptors.

scholarly journals Cell transformation by fibroblast growth factors can be suppressed by truncated fibroblast growth factor receptors.

1994 ◽  
Vol 14 (11) ◽  
pp. 7660-7669 ◽  
Author(s):  
Y Li ◽  
C Basilico ◽  
A Mansukhani
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Tyrosine Kinases ◽  
Dominant Negative ◽  
Receptor Expression ◽  
Scatchard Analysis ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Truncated Receptor ◽  
Nih 3T3

Ligand-induced dimerization and transphosphorylation are thought to be important events by which receptor tyrosine kinases generate cellular signals. We have investigated the ability of signalling-defective, truncated fibroblast growth factor (FGF) receptors (FGFR-1 and FGFR-2) to block the FGF response in cells that express both types of endogenous FGF receptors. When these dominant negative receptors are expressed in NIH 3T3 cells transformed by the secreted FGF-4, the transformed properties of the cells can be reverted to various degrees, with better reversion phenotype correlating with higher levels of truncated receptor expression. Furthermore, truncated FGFR-2 is significantly more efficient at producing reversion than FGFR-1, indicating that FGF-4 preferentially utilizes the FGFR-2 signalling pathway. NIH 3T3 clones expressing these truncated receptors are more resistant to FGF-induced mitogenesis and also exhibit reduced tyrosine phosphorylation upon treatment with FGF. The block in FGF-signalling, however, can be overcome by the addition of excess growth factor. The truncated receptors have binding affinities that are four- to eightfold lower than those of wild-type receptors, as measured by Scatchard analysis. We also observed a partial specificity in the responses of truncated-receptor-expressing clones to FGF-2 or FGF-4. Our results suggest that the block to signal transduction produced by kinase-negative FGF receptors is achieved through a combination of dominant negative effects and competition for growth factor binding with functional receptors.

scholarly journals Self-renewal of human embryonic stem cells requires insulin-like growth factor-1 receptor and ERBB2 receptor signaling

Blood ◽  
2007 ◽  
Vol 110 (12) ◽  
pp. 4111-4119 ◽  
Author(s):  
Linlin Wang ◽  
Thomas C. Schulz ◽  
Eric S. Sherrer ◽  
Derek S. Dauphin ◽  
Soojung Shin ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Tyrosine Phosphorylation ◽  
Tyrosine Kinases ◽  
Embryonic Stem ◽  
Growth Factor Receptor ◽  
Insulin Like Growth Factor ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Self Renewal

Abstract Despite progress in developing defined conditions for human embryonic stem cell (hESC) cultures, little is known about the cell-surface receptors that are activated under conditions supportive of hESC self-renewal. A simultaneous interrogation of 42 receptor tyrosine kinases (RTKs) in hESCs following stimulation with mouse embryonic fibroblast (MEF) conditioned medium (CM) revealed rapid and prominent tyrosine phosphorylation of insulin receptor (IR) and insulin-like growth factor-1 receptor (IGF1R); less prominent tyrosine phosphorylation of epidermal growth factor receptor (EGFR) family members, including ERBB2 and ERBB3; and trace phosphorylation of fibroblast growth factor receptors. Intense IGF1R and IR phosphorylation occurred in the absence of MEF conditioning (NCM) and was attributable to high concentrations of insulin in the proprietary KnockOut Serum Replacer (KSR). Inhibition of IGF1R using a blocking antibody or lentivirus-delivered shRNA reduced hESC self-renewal and promoted differentiation, while disruption of ERBB2 signaling with the selective inhibitor AG825 severely inhibited hESC proliferation and promoted apoptosis. A simple defined medium containing an IGF1 analog, heregulin-1β (a ligand for ERBB2/ERBB3), fibroblast growth factor-2 (FGF2), and activin A supported long-term growth of multiple hESC lines. These studies identify previously unappreciated RTKs that support hESC proliferation and self-renewal, and provide a rationally designed medium for the growth and maintenance of pluripotent hESCs.

scholarly journals Fibroblast growth factor receptors in breast cancer

Tumor Biology ◽  
2017 ◽  
Vol 39 (5) ◽  
pp. 101042831769837 ◽  
Author(s):  
Shuwei Wang ◽  
Zhongyang Ding
Keyword(s):  
Breast Cancer ◽  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Tyrosine Kinases ◽  
Growth Factor Receptor ◽  
Growth Factor Receptors ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Genetic Studies ◽  
The Common

Fibroblast growth factor receptors are growth factor receptor tyrosine kinases, exerting their roles in embryogenesis, tissue homeostasis, and development of breast cancer. Recent genetic studies have identified some subtypes of fibroblast growth factor receptors as strong genetic loci associated with breast cancer. In this article, we review the recent epidemiological findings and experiment results of fibroblast growth factor receptors in breast cancer. First, we summarized the structure and physiological function of fibroblast growth factor receptors in humans. Then, we discussed the common genetic variations in fibroblast growth factor receptors that affect breast cancer risk. In addition, we also introduced the potential roles of each fibroblast growth factor receptors isoform in breast cancer. Finally, we explored the potential therapeutics targeting fibroblast growth factor receptors for breast cancer. Based on the biological mechanisms of fibroblast growth factor receptors leading to the pathogenesis in breast cancer, targeting fibroblast growth factor receptors may provide new opportunities for breast cancer therapeutic strategies.

scholarly journals Fibroblast Growth Factor Receptor Functions in Glioblastoma

Cells ◽  
2019 ◽  
Vol 8 (7) ◽  
pp. 715 ◽  
Author(s):  
Ana Jimenez-Pascual ◽  
Florian A. Siebzehnrubl
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor ◽  
Tyrosine Kinases ◽  
Therapeutic Potential ◽  
Growth Factor Receptor ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors ◽  
Comprehensive Overview ◽  
Tyrosine Kinase Signaling ◽  
Molecular Alterations

Glioblastoma is the most lethal brain cancer in adults, with no known cure. This cancer is characterized by a pronounced genetic heterogeneity, but aberrant activation of receptor tyrosine kinase signaling is among the most frequent molecular alterations in glioblastoma. Somatic mutations of fibroblast growth factor receptors (FGFRs) are rare in these cancers, but many studies have documented that signaling through FGFRs impacts glioblastoma progression and patient survival. Small-molecule inhibitors of FGFR tyrosine kinases are currently being trialed, underlining the therapeutic potential of blocking this signaling pathway. Nevertheless, a comprehensive overview of the state of the art of the literature on FGFRs in glioblastoma is lacking. Here, we review the evidence for the biological functions of FGFRs in glioblastoma, as well as pharmacological approaches to targeting these receptors.

scholarly journals Cross-Talk between Fibroblast Growth Factor Receptors and Other Cell Surface Proteins

Cells ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 455 ◽  
Author(s):  
Marta Latko ◽  
Aleksandra Czyrek ◽  
Natalia Porębska ◽  
Marika Kucińska ◽  
Jacek Otlewski ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Cell Fate ◽  
Cross Talk ◽  
Tyrosine Kinases ◽  
Intracellular Signaling ◽  
Cellular Response ◽  
Surface Proteins ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors

Fibroblast growth factors (FGFs) and their receptors (FGFRs) constitute signaling circuits that transmit signals across the plasma membrane, regulating pivotal cellular processes like differentiation, migration, proliferation, and apoptosis. The malfunction of FGFs/FGFRs signaling axis is observed in numerous developmental and metabolic disorders, and in various tumors. The large diversity of FGFs/FGFRs functions is attributed to a great complexity in the regulation of FGFs/FGFRs-dependent signaling cascades. The function of FGFRs is modulated at several levels, including gene expression, alternative splicing, posttranslational modifications, and protein trafficking. One of the emerging ways to adjust FGFRs activity is through formation of complexes with other integral proteins of the cell membrane. These proteins may act as coreceptors, modulating binding of FGFs to FGFRs and defining specificity of elicited cellular response. FGFRs may interact with other cell surface receptors, like G-protein-coupled receptors (GPCRs) or receptor tyrosine kinases (RTKs). The cross-talk between various receptors modulates the strength and specificity of intracellular signaling and cell fate. At the cell surface FGFRs can assemble into large complexes involving various cell adhesion molecules (CAMs). The interplay between FGFRs and CAMs affects cell–cell interaction and motility and is especially important for development of the central nervous system. This review summarizes current stage of knowledge about the regulation of FGFRs by the plasma membrane-embedded partner proteins and highlights the importance of FGFRs-containing membrane complexes in pathological conditions, including cancer.

scholarly journals The Acidic Domain and First Immunoglobulin-Like Loop of Fibroblast Growth Factor Receptor 2 Modulate Downstream Signaling through Glycosaminoglycan Modification

1999 ◽  
Vol 19 (10) ◽  
pp. 6754-6764 ◽  
Author(s):  
Kazushige Sakaguchi ◽  
Matthew V. Lorenzi ◽  
Donald P. Bottaro ◽  
Toru Miki
Keyword(s):  
Growth Factor ◽  
Heparan Sulfate ◽  
Fibroblast Growth Factor ◽  
Tyrosine Kinases ◽  
Mitogen Activated Protein Kinase ◽  
Alternative Exon ◽  
Protein Kinase Activity ◽  
Regulation Mechanism ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors

ABSTRACT Fibroblast growth factor receptors (FGFRs) are membrane-spanning tyrosine kinases that have been implicated in a variety of biological processes including mitogenesis, cell migration, development, and differentiation. We identified a unique isoform of FGFR2 expressed as a diffuse band with an unusually large molecular mass. This receptor is modified by glycosaminoglycan at a Ser residue located immediately N terminal to the acidic box, a stretch of acidic amino acids. The acidic box and the glycosaminoglycan modification site are encoded by an alternative exon of the FGFR2 gene. The acidic box appears to play an important role in glycosaminoglycan modification, and the presence of this domain is required for modification by heparan sulfate glycosaminoglycan. Moreover, the presence of the first immunoglobulin-like domain encoded by another alternative exon abrogated the modification. The high-affinity receptor with heparan sulfate modification enhanced receptor autophosphorylation, substrate phosphorylation, and ternary complex factor-independent gene expression. It also sustained mitogen-activated protein kinase activity and increased eventual DNA synthesis, a long-term response to fibroblast growth factor stimulation, at physiological ligand concentrations. We propose a novel regulation mechanism of FGFR2 signal transduction through glycosaminoglycan modification.

scholarly journals Fibroblast Growth Factor Receptors (FGFRs): Structures and Small Molecule Inhibitors

Cells ◽  
2019 ◽  
Vol 8 (6) ◽  
pp. 614 ◽  
Author(s):  
Shuyan Dai ◽  
Zhan Zhou ◽  
Zhuchu Chen ◽  
Guangyu Xu ◽  
Yongheng Chen
Keyword(s):  
Growth Factor ◽  
Urothelial Carcinoma ◽  
Fibroblast Growth Factor ◽  
Small Molecule ◽  
Tyrosine Kinases ◽  
Drug Targets ◽  
Small Molecule Inhibitors ◽  
Growth Factor Receptors ◽  
Fibroblast Growth ◽  
Fibroblast Growth Factor Receptors

Fibroblast growth factor receptors (FGFRs) are a family of receptor tyrosine kinases expressed on the cell membrane that play crucial roles in both developmental and adult cells. Dysregulation of FGFRs has been implicated in a wide variety of cancers, such as urothelial carcinoma, hepatocellular carcinoma, ovarian cancer and lung adenocarcinoma. Due to their functional importance, FGFRs have been considered as promising drug targets for the therapy of various cancers. Multiple small molecule inhibitors targeting this family of kinases have been developed, and some of them are in clinical trials. Furthermore, the pan-FGFR inhibitor erdafitinib (JNJ-42756493) has recently been approved by the U.S. Food and Drug Administration (FDA) for the treatment of metastatic or unresectable urothelial carcinoma (mUC). This review summarizes the structure of FGFR, especially its kinase domain, and the development of small molecule FGFR inhibitors.

Sign in / Sign up

Export Citation Format

Share Document