90-kDa Ribosomal S6 Kinase Is a Direct Target for the Nuclear Fibroblast Growth Factor Receptor 1 (FGFR1)

Abstract

Fibroblast growth factor receptor 1 (FGFR1) is a transmembrane protein capable of transducing stimulation by secreted FGFs. In addition, newly synthesized FGFR1 enters the nucleus in response to cellular stimulation and during development. Nuclear FGFR1 can transactivate CRE (cAMP responsive element), activate CRE-binding protein (CREB)-binding protein (CBP) and gene activities causing cellular growth and differentiation. Here, a yeast two-hybrid assay was performed to identify FGFR1-binding proteins and the mechanism of nuclear FGFR1 action. Ten FGFR1-binding proteins were identified. Among the proteins detected with the intracellular FGFR1 domain was a 90-kDa ribosomal S6 kinase (RSK1), a regulator of CREB, CBP, and histone phosphorylation. FGFR1 bound to the N-terminal region of RSK1. The FGFR1-RSK1 interaction was confirmed by co-immunoprecipitation and colocalization in the nucleus and cytoplasm of mammalian cells. Predominantly nuclear FGFR1-RSK1 interaction was observed in the rat brain during neurogenesis and in cAMP-stimulated cultured neural cells. In TE671 cells, transfected FGFR1 colocalized and coimmunoprecipitated, almost exclusively, with nuclear RSK1. Nuclear RSK1 kinase activity and RSK1 activation of CREB were enhanced by transfected FGFR1. In contrast, kinase-deleted FGFR1 (TK؊), which did not bind to RSK1 failed to stimulate nuclear RSK1 activity or RSK1 activation of CREB. Kinase inactive FGFR1 (K514A) bound effectively to nuclear RSK1, but it failed to stimulate RSK1. Thus, active FGFR1 kinase regulates the functions of nuclear RSK1. The interaction of nuclear FGFR1 with pluripotent RSK1 offers a new mechanism through which FGFR1 may control fundamental cellular processes.Fibroblast growth factor receptor 1 (FGFR1), 1 like other single transmembrane growth factor receptors, transduces signals across the cell membrane generated by extracellular ligands (1). Binding of secreted FGFs causes FGFR1 molecules to dimerize and undergo cross-phosphorylation (2, 3). The phosphotyrosine residues serve as docking sites for the SH2 (Src homology 2) or phosphotyrosine-binding domains of cytoplasmic proteins, which initiate downstream signaling cascades. A number of proteins that interact with the cytosolic, C-terminal portion of FGFR1 have been identified, including phospholipase C␥ (3, 4), adaptor proteins Shc (5), Grb14 (6), the regulatory subunit (p85) of the phosphatidylinositol 3Ј-kinase (PI3K), and NCK (7). However, binding of the FGF receptor substrate 2 to FGFR1 occurs independent of ligand stimulation and tyrosine phosphorylation (8, 9), suggesting multiple mechanisms of FGFR1 action.Studies in this and other laboratories have shown that upon cell stimulation, FGFR1, a typically plasma membrane-associated protein, translocates to the cell nucleus along with nonsecreted, intracellular forms of FGF-2, which lack a signal peptide but contain a functional nuclear localization signal (NLS) (10). Nuclear FGFR1 is full-length, binds FGF-2, and has an active TK domain (11,12). Nuclear FGFR1 is not deriv...

Organically modified silica nanoparticles: A nonviral vector for in vivo gene delivery and expression in the brain

“…To be able to therapeutically manipulate the endogenous adult NSPCs in the brain SVZ, it is crucial to identify the extracellular and intracellular signals that regulate division and control the fate of these cells (44,45). In cultured cells and in the developing rat brain, FGFR1 was associated with the peripheral cytoplasm but also with the cell nuclei (46,47). The cell-surface FGFR mediates the mitogenic effects of extracellular FGFs, whereas the nonmembrane nuclear FGFR1 signals withdrawal from the cell cycle and postmitotic growth (46).…”

Fibroblast Growth Factor Receptor-1 (FGFR1) Nuclear Dynamics Reveal a Novel Mechanism in Transcription Control

“…The FGFR1(TKϪ) mutant binds to CBP, but lacks the tyrosine kinase (TK) domain that is essential to activate transcription (Stachowiak et al, 2003b;Fang et al, 2005) and to bind to many other proteins (Hu et al, 2004;Fang et al, 2005). Transfected FGFR1(TKϪ)-EGFP displayed an immobile nuclear population that was twice as large (72.68%; Table 1) as the full-length FGFR1-EGFP (36.18%).…”

“…These events are accompanied by the nuclear coaccumulation of FGF-2, FGFR1 binding to the transcription coactivator CBP, and transcriptional upregulation (Stachowiak et al, 2003b;Hu et al, 2004;Fang et al, 2005).…”

“…Human TE671 cells and neuroblastoma BE(2)C ( Lee and Kim, 2004) were cultured and transfected as previously (Hu et al, 2004;Fang et al, 2005;Dunham-Ems et al, 2006). The efficiency of TE671 transfection was greater than 70% as in our previous studies.…”

Exogenously Added Fibroblast Growth Factor 2 (FGF-2) to NIH3T3 CellsInteracts with Nuclear Ribosomal S6 Kinase 2 (RSK2) in a Cell Cycle-dependentManner

“…Recently, Hu et al, have isolated RSK1⅐FGFR1 complexes from the nuclei of the prenatal rat brain (33). RSK1 and FGFR1 were never detected in FGF-2⅐RSK2 complexes in vivo (data not shown), and in vitro RSK1 does not interact with FGF-2.…”