Alternative splicing in fibroblast growth factor receptor 2 is associated with induced epithelial-mesenchymal transition in rat bladder carcinoma cells.

Abstract

We described previously that acidic fibroblast growth factor (aFGF), but not basic fibroblast growth factor (bFGF), can induce the rat carcinoma cell line NBT-II to undergo a rapid and reversible transition from epithelial to mesenchymal phenotype (EMT). We now find that NBT-II EMT is stimulated by keratinocyte growth factor (KGF) in cells grown at low density. Accordingly, a high-affinity receptor showing 98% homology to mouse FGF receptor 2b/KGF receptor was cloned and sequenced from NBT-II cells. Northern analysis indicated that mRNA for FGF receptor 2b/KGF receptor was drastically down-regulated within 1 wk in aFGF-induced mesenchymal NBT-II cells. This decrease coincided with an up-regulation of FGF receptor 2c/Bek, a KGF-insensitive, alternatively spliced form of FGF receptor 2b/KGF receptor. Functional studies confirmed that KGF could not maintain EMT induction on mesenchymal NBT-II cells. FGF receptor 1 and FGF receptor 2c/Bek could also support EMT induction when transfected into NBT-II cells in response to aFGF or bFGF. Such transfected cells could bind bFGF as well as aFGF. Therefore, EMT can be induced through different FGF receptors, but EMT may also regulate FGF receptor expression itself.

Exon III splicing switch of fibroblast growth factor (FGF) receptor-2 and -3 can be induced by FGF-1 or FGF-2

“…We observed with the two cell lines induced to produce endogenous FGF upon transfection an oscillation between a mixture of IIIb and IIIc, when the cells were subcon¯uent and only IIIb transcripts when the cells returned to con¯uency. In contradiction with others (Savagner et al, 1994), we did not observe that the cell con¯uency per se, without adding FGF, induce an exon III switch. Con¯uent NBT-II or SVK14 cells, as subcon¯uent cells, showed only IIIb transcripts.…”

“…It was also demonstrated by this group that NBT-II cells activated for 1 week with FGF-1 switched from FGFR-2/IIIb to FGFR-2/IIIc (Savagner et al, 1994). However, as NBT-II cells cultured at high density, without FGF-1, also switched to IIIc (Savagner et al, 1994), it was not clear from this study if the FGF receptor exon switching could be induced by cell con¯uency as well as by FGF-1. This prompted us to reexamine the in¯uence of FGFs and of cell con¯uency on the IIIb/IIIc choice in the FGFR genes.…”

The community effect in FGF-1 mediated tumor progression of a rat bladder carcinoma does not involve a direct paracrine signaling

“…Cross-linking experiments with iodinated FGF-1 (Figure 2), clearly show the presence of the NBT-II endogeneous FGF receptors which have been previously characterized as the KGFR/FGFR2b alternatively spliced variant of the FGFR2 receptor (Savagner et al, 1994). Iodinated FGF-1 can be totally displaced by competition with addition of cold FGF-1 (106concentration) but not by addition of FGF-2.…”

“…These epithelial cells do not produce either FGF-1 or FGF-2 but possess high a nity receptors for FGF-1 (Valles et al, 1990) which was cloned and identi®ed as the FGFR2b spliced variant of the FGFR2 receptor (ie KGFR) (Savagner et al, 1994).…”

“…Transfected NBT-II cells expressing FGF-1 (NSF14 cells) or the FGFR1 (Flg receptor) have been reported elsewhere (Jouanneau et al, 1991;Savagner et al, 1994) MDCK cells (dog kidney epithelial cells) are a kind gift from Dr R Warn (University of East Anglia, Norwich, England) and are routinely used in our laboratory to test the scattering activity of the Scatter Factor (HGF/SF) produced either in the supernatant of MRC5 naturally producing ®broblastic cells or present in the conditioned medium of HGF/SF transfected NBT-II cells (Bellusci et al, 1994).…”