Two Nuclear Localization Signals Required for Transport from the Cytosol to the Nucleus of Externally Added FGF-1 Translocated into Cells

Abstract

Externally added FGF-1 is transported into the nucleus of cells. It was earlier shown that FGF-1 contains an N-terminal nuclear localization signal (NLS) implicated in the stimulation of DNA synthesis. We here provide evidence that FGF-1 contains a second putative NLS (NLS2), which is located near the C-terminus. It is a bipartite NLS consisting of two clusters of lysines separated by a spacer of 10 amino acids. A fusion protein of GFP and the bipartite NLS was more efficiently transported into the nucleus than GFP alone, indicating that it can act as an NLS in the living cell. FGF-1 mutated in the N-terminal NLS (NLS1) or in the first cluster of the bipartite NLS2 bound to heparin and FGF receptors and activated downstream signaling similarly to the wild-type growth factor. Mutations in the second cluster of NLS2 resulted in impaired interaction with heparin and reduced stability. When radiolabeled FGF-1 with mutated NLS1 or the first lysine cluster of NLS2 was added to NIH/3T3 cells, it was translocated into the cytosol, but not transported efficiently to the nucleus. Phosphorylation of FGF-1 occurs normally in the nucleus, and while wild-type FGF-1 was phosphorylated after addition to cells, the NLS mutants were not. It therefore appears that both NLS1 and NLS2 are important for efficient transport of FGF-1 to the nucleus. Stimulation of DNA synthesis by FGF-1 with mutations in both NLSs was reduced considerably indicating that efficient transport to the nucleus may be involved in the stimulation of DNA synthesis.

Characterization of FN1–FGFR1 and novel FN1–FGF1 fusion genes in a large series of phosphaturic mesenchymal tumors

“…24,25 The expected retention of the nuclear localization sequence of FGF1 in this fusion protein is noteworthy, as the receptormediated internalization and nuclear localization sequence-dependent nuclear translocation of FGF1 is crucial for FGF1-induced mitogenic effects. 26,27 Of interest, focal nuclear staining of FGF1 was observed in some phosphaturic mesenchymal tumors regardless of the fusion status, suggesting that the biological functions of nuclear FGF1 might be in effect in these tumors.…”

Localization of Fibroblast Growth Factor-1 in Cholinergic Neurons Innervating the Rat Larynx

“…Some isoforms of FGF1 receptors exist in the perinuclear region of the cell body (Prudovsky et al 1996). In addition, FGF1 contains nuclear localization signals and plays a role in nuclear functioning (Imamura et al 1990;Wesche et al 2005). The predominance of FGF1 in cell bodies may therefore result from interaction with these receptors in the perinuclear region.…”

Different abilities of the four FGFRs to mediate FGF-1 translocation are linked to differences in the receptor C-terminal tail

“…Intracellular FGF-2 has been shown to interact directly with upstream binding factor (UBF) and stimulate rRNA transcription (Sheng et al, 2005). Nuclear import is directed by two nuclear localization sequences in FGF-1 (Imamura et al, 1990;Wiedlocha et al, 1994;Imamura et al, 1994;Friedman et al, 1994;Wesche et al, 2005). The translocated FGF-1 is phosphorylated at a single site, at S130, by PKC␦.…”