Modulation of cell growth and transformation by doxycycline-regulated FGF-2 expression in NIH-3T3 cells.
The single-copy fibroblast growth factor 2 (FGF-2) gene encodes four coexpressed isoforms of different molecular masses. The 18-kDa FGF-2 is primarily localized in the cytoplasm, whereas the higher molecular mass isoforms (HMW FGF-2) localize to the nucleus and nucleolus. The overexpression of either 18-kDa FGF-2 or HMW FGF-2 promotes cell transformation in a dose-dependent manner. In NIH 3T3 cells, the selective overexpression of HMW FGF-2 but not of 18-kDa FGF-2 confers upon the cells the unique phenotype of growth in low serum-containing medium. Thus, the distinct intracellular localization and the level of expression of FGF-2 are pivotal requirements for the differential effects of FGF-2 isoforms on the cellular phenotype. On this basis, we established a doxycycline-regulatable FGF-2 expression system that permitted us to regulate the expression of each isoform in a time- and dose-dependent manner. We analyzed the growth properties of cells in the presence and absence of doxycycline in both normal and low serum-containing medium and in soft agar. The doxycycline-activated expression of 18-kDa FGF-2 did not allow growth in low serum medium. The growth of cells expressing HMW FGF-2 was increased by doxycycline under all three conditions, and a relationship between the level of HMW FGF-2 expression and cell growth was observed for all three conditions. This doxycycline-regulatable FGF-2 expression system provides a mechanism to analyze changes in FGF-2 targeted pathways and genes and to characterize pathways specifically activated by either the 18-kDa FGF-2 or the HMW FGF-2 isoforms.
Gualandris
A
A
Department of Cell Biology, New York University Medical Center, New York, New York, USA. gualaa01@mcrcr.med.nyu.edu
Arese
M
M
Shen
B
B
Rifkin
D B
DB
D016475:3T3 Cells; D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002461:Cell Line, Transformed; D002999:Clone Cells; D004305:Dose-Response Relationship, Drug; D004318:Doxycycline / Q000494:pharmacology*; D016222:Fibroblast Growth Factor 2 / Q000235:genetics* / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D051379:Mice; D020033:Protein Isoforms / Q000235:genetics; D014162:Transfection
D020033:Protein Isoforms; D016222:Fibroblast Growth Factor 2; D004318:Doxycycline