Cloning of an intracellular protein that binds selectively to mitogenic acidic fibroblast growth factor.
In addition to its extracellular action, there is evidence that acidic fibroblast growth factor (aFGF) acts inside cells. To identify intracellular proteins interacting with aFGF, we screened a HeLa cell library in the yeast two-hybrid system using pLex-aFGF as a bait. A clone binding to aFGF, but not to the non-mitogenic mutant aFGF-K132E, was isolated and characterized. The insert contained an open reading frame corresponding to a novel protein of 42 kDa. The protein, termed aFGF intracellular binding protein (FIBP), is mainly hydrophilic and does not contain an N-terminal signal sequence. In vitro-translated FIBP bound specifically to a fusion protein of maltose-binding protein and aFGF. FIBP became post-translationally associated with microsomes added to the cell-free protein synthesizing system, and the membrane-associated protein bound aFGF with high efficiency. Immunoblots and fluorescence microscopy demonstrated that the protein is present in nuclei and, to a lesser extent, associated with mitochondria and other cytoplasmic membranes. The possibility is discussed that FIBP may be involved in the mitogenic action of aFGF.
Kolpakova
E
E
Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, 0310 Oslo, Norway.
Wiedłocha
A
A
Stenmark
H
H
Klingenberg
O
O
Falnes
P O
PO
Olsnes
S
S
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D004261:DNA Replication; D018076:DNA, Complementary; D016220:Fibroblast Growth Factor 1 / Q000235:genetics / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008934:Mitogens / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000378:metabolism; D014709:Vero Cells
D002352:Carrier Proteins; D018076:DNA, Complementary; C116274:FIBP protein, human; D008565:Membrane Proteins; D008934:Mitogens; D012333:RNA, Messenger; D016220:Fibroblast Growth Factor 1