cDNA Cloning and Expression of a Human FGF Receptor which Binds Acidic and Basic FGF

Abstract

We have isolated and characterized a cDNA clone, phFGFR, encoding a human fibroblast growth factor (FGF) receptor. phFGFR contains an open reading frame which encodes an 820 amino acid polypeptide with three immunoglobulin-like domains in the extracellular part and an intracellular split tyrosine kinase domain. Transient expression in COS-1 cells and immunoprecipitation using an antiserum raised against a C-terminal peptide, gave rise to two components, representing mature (130 kDa) and precursor (115 kDa) forms of the phFGFR encoded polypeptide, which was denoted hFGFR-1. Crosslinking of iodinated acidic FGF (aFGF) and basic FGF (bFGF) to transiently expressing COS-1 cells revealed a major band of 95 kDa, which was competed for by both aFGF and bFGF. From Scatchard analyses, the Kd:s for binding of aFGF and bFGF to hFGFR-1 were estimated to 25 pM and 41 pM, respectively. Thus, phFGFR encodes a human FGF receptor with high affinity for both aFGF and bFGF.

Shc and a Novel 89-kDa Component Couple to the Grb2-Sos Complex in Fibroblast Growth Factor-2-stimulated Cells

“…Thus far, two such adaptor molecules have been thoroughly characterized, Shc (33) and PTP 1D/Syp (27). We could not detect direct interaction between FGFR-1 and Grb2, which is consistent with the lack of consensus binding site for Grb2, Y(P)-X-N, in the FGFR-1 sequence (34). We further showed that PTP 1D/Syp is not tyrosine-phosporylated in FGF-2-stimulated cells.…”

Putative tyrosine kinases expressed in K-562 human leukemia cells.

“…1B shows the homology of JTK2 and JTK4 sequences to other members of the FGFR family. These include human FLG (hFGFR-1), originally cloned from a human endothelial cell cDNA library (31)(32)(33), its mouse homologue (34), and the corresponding chicken gene cek-J (39,40) and another member of the FGFR family, the mouse bek gene (41). The presence of several FGFRrelated mRNA sequences in the K-562 cells is particularly exciting, as there is also a gene family encoding several different fibroblast growth factors (35)(36)(37)(42)(43)(44)(45)(46).…”

Different intracellular trafficking of FGF1 endocytosed by the four homologous FGF receptors

“…Cy3-maleimide, heparin-Sepharose, streptavidin-Sepharose, ECL plus western blotting system were from Amersham Biosciences (Buckinghamshire, UK Plasmids pcDNA3-hFGFR1: cDNA encoding hFGFR1 IIIc was cut out from pSV7d (Wennstrom et al, 1991) with EcoRI and XbaI in two fragments, and ligated into pcDNA3 (Invitrogen, Carlsbad, CA) cut with the same enzymes. pcDNA3-hFGFR2: cDNA encoding hFGFR2 IIIc lacking D1 was cut out from pBluescript (RZPD, Berlin, Germany, Clone ID: IMAGp998N0911701Q3) with NotI and SpeI, and ligated into pcDNA3 cut with NotI and XbaI.…”