Proteomic detection of changes in protein synthesis induced by fibroblast growth factor-2 in MCF-7 human breast cancer cells.
Fibroblast growth factor-2 (FGF-2) is a potent regulator of breast cancer cell growth through stimulation of tyrosine kinase receptors and activation of the mitogen-activated protein kinase cascade. In the present study, we have investigated changes in protein synthesis induced by FGF-2 stimulation of the prototypic human breast cancer cell line MCF-7. Using high-resolution two-dimensional electrophoresis of (35)S amino acid metabolically labeled proteins and computerized analysis of 2D autoradiograms, we found that four proteins were up-regulated within the first 12 h of FGF-2 stimulation. Mass spectrometry analysis (MALDI-TOF and MS-MS) of tryptic fragments and database searches allowed the identification of these FGF-2-regulated proteins as the heat shock proteins HSP90 and HSP70, the proliferating cell nuclear antigen (PCNA), and the transcriptionaly controlled tumor protein (TCTP). We then analyzed the distribution of these proteins in various cancerous and normal breast epithelial cells. Interestingly, the four FGF-2-regulated proteins were found to be constitutively up-regulated in ras-transfected MCF-7 cells, indicating their relevance to the up-regulation of cellular proliferation. Moreover, HSP90 and PCNA were found at higher levels in cancerous cells than in normal cells. The role of HSP90 was further investigated using the specific inhibitor geldanamycin. We showed that the functionality of HSP90 is strictly required in order to obtain FGF-2 mitogenic stimulation in MCF-7 cells, indicating the crucial role played by this molecular chaperone in the control of breast cancer cell growth. Finally, these results show that proteomic analysis is a valuable method for identifying potential markers or therapeutic targets related to cancer growth.
Vercoutter-Edouart
A S
AS
Equipe Facteurs de Croissance, UMR 8576 CNRS, Villeneuve d'Ascq, France.
Czeszak
X
X
Crépin
M
M
Lemoine
J
J
Boilly
B
B
Le Bourhis
X
X
Peyrat
J P
JP
Hondermarck
H
H
D014408:Biomarkers, Tumor; D001943:Breast Neoplasms; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002455:Cell Division; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism* / Q000494:pharmacology; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D020543:Proteome / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D014407:Tumor Cells, Cultured; D000090324:Tumor Protein, Translationally-Controlled 1
D014408:Biomarkers, Tumor; D002135:Calcium-Binding Proteins; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D018809:Proliferating Cell Nuclear Antigen; D020543:Proteome; C000715647:TPT1 protein, human; D000090324:Tumor Protein, Translationally-Controlled 1; D016222:Fibroblast Growth Factor 2