Translocation of cytosol of exogenous, CAAX-tagged acidic fibroblast growth factor.
Acidic fibroblast growth factor (aFGF) added externally to cells has been proposed to enter the nucleus and stimulate DNA synthesis, but it has remained controversial whether or not exogenous aFGF has the capability of crossing cellular membranes. To test this, a novel principle to study translocation of proteins to the cytosol was developed by fusing a C-terminal farnesylation signal, a CAAX tag (C = Cys, A = an aliphatic amino acid, and X = any amino acid), onto aFGF. Farnesylation is only known to occur in the cytosol and possibly in the nucleus. When incubated with NIH3T3 cells overnight, about one-third of the cell-associated, CAAX-tagged growth factor was farnesylated, indicating that efficient translocation had taken place. Binding to specific FGF receptors was required for translocation to occur. Part of the farnesylated growth factor was found in the nuclear fraction. The data indicate that CAAX-tagged aFGF added externally to cells is able to cross cellular membranes and enter the cytosol and the nucleus.
Wiedłocha
A
A
Institute for Cancer Research, Norwegian Radium Hospital, Montebello, Oslo, Norway.
Falnes
P O
PO
Rapak
A
A
Klingenberg
O
O
Muñoz
R
R
Olsnes
S
S
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D016220:Fibroblast Growth Factor 1 / Q000096:biosynthesis / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D012516:Osteosarcoma; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017468:Receptors, Fibroblast Growth Factor / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism / Q000494:pharmacology; D016324:Sequence Tagged Sites; D012717:Sesquiterpenes; D014162:Transfection; D014407:Tumor Cells, Cultured
D011106:Polyisoprenyl Phosphates; D017468:Receptors, Fibroblast Growth Factor; D011994:Recombinant Proteins; D012717:Sesquiterpenes; D016220:Fibroblast Growth Factor 1; C004808:farnesyl pyrophosphate