Quantitative export of FGF-2 occurs through an alternative, energy-dependent, non-ER/Golgi pathway.
Although basic fibroblast growth factor (bFGF/FGF-2) is found outside cells, it lacks a conventional signal peptide sequence; the mechanism underlying its export from cells is therefore unknown. Using a transient COS-1 cell expression system, we have identified a novel membrane-associated transport pathway that mediates export of FGF-2. This export pathway is specific for the 18-kD isoform of FGF-2, is resistant to the anti-Golgi effects of Brefeldin A, and is energy-dependent. In FGF-2-transfected COS-1 cells, this ER/Golgi-independent pathway appears to be constitutively active and functions to quantitatively export metabolically-labeled 18-kD FGF-2. Co-transfection and co-immunoprecipitation experiments, using a vector encoding the cytoplasmic protein neomycin phosphotransferase, further demonstrated the selectivity of this export pathway for FGF-2. When neomycin phosphotransferase was appended to the COOH-terminus of 18-kD FGF-2, the chimera was exported. However, the transmembrane anchor sequence of the integral membrane glycoprotein (G protein) of vesicular stomatitis virus (VSV) blocked export. The chimeric protein localized to the plasma membrane with its FGF-2 domain extracellular and remained cell-associated following alkaline carbonate extraction. Taken together, the data suggest that FGF-2 is "exported" from cells via a unique cellular pathway, which is clearly distinct from classical signal peptide-mediated secretion. This model system provides a basis for the development and testing of therapeutic agents which may block FGF-2 export. Such an intervention may be of considerable use for the treatment of angiogenesis-dependent diseases involving FGF-2.
Department of Molecular and Cellular Growth Biology, Whittier Institute, La Jolla, California 92037.
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RZ
Florkiewicz
R A
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Majack
R D
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Buechler
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Florkiewicz
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001693:Biological Transport, Active; D020126:Brefeldin A; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003517:Cyclopentanes / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D019868:Kanamycin Kinase; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000378:metabolism
D003517:Cyclopentanes; D017931:DNA Primers; C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; D016222:Fibroblast Growth Factor 2; D020126:Brefeldin A; D017853:Phosphotransferases (Alcohol Group Acceptor); D019868:Kanamycin Kinase