Insulin-like growth factor binding protein-3 induces G1 cell cycle arrest with inhibition of cyclin-dependent kinase 2 and 4 in MCF-7 human breast cancer cells.
Insulin-like growth factor binding protein (IGFBP)-3 has been shown to potently inhibit proliferation of various cell types in an insulin-like growth factor (IGF)-independent manner. We have previously shown that IGFBP-3 induces apoptosis in an IGF-independent manner through the activation of caspases involved in a death receptor-mediated pathway in MCF-7 human breast cancer cells. In the present study, we present further evidence that IGFBP-3 inhibits cell proliferation through the induction of cell cycle arrest in the same cell line. Induction of IGFBP-3 in MCF-7 cells inhibited cell proliferation whereas presence of small interfering RNA against IGFBP-3 abolished cell inhibitory effect of IGFBP-3, suggesting that the observed growth inhibition is specific. Flow cytometry analysis showed that induced expression of IGFBP-3 led to an arrest of the cell cycle in G1-S phase. Western immunoblot analysis showed a significant decrease in the levels of the cell cycle-regulated proteins such as cyclin D1, cyclin D3, cyclin E, cyclin A, cyclin-dependent kinase (CDK) 2, CDK4, retinoblastoma protein (pRB), and phosph-pRB, suggesting a possible mechanism for cell cycle arrest by IGFBP-3. Northern blot analysis and real-time quantitative PCR demonstrated a significant decrease in gene expression of cyclin D1. Additional phosphorylation assay showed that IGFBP-3 decreased the phosphorylation activity of CDK2 and CDK4. These results show that cellular production of IGFBP-3 leads to G1 cell cycle arrest with inhibition of CDK2 and CDK4. Taken together, IGFBP-3 exerts its growth inhibitory action through not only induction of apoptosis but also the G1 cell cycle arrest in human breast cancer cells.
Kim
H-S
HS
Department of Pediatrics, Yonsei University, Seoul, Korea. kimho@yuhs.ac
Lee
W J
WJ
Lee
S W
SW
Chae
H-W
HW
Kim
D H
DH
Oh
Y
Y
D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051358:Cyclin-Dependent Kinase 4 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D050756:Cyclin-Dependent Kinase Inhibitor Proteins / Q000378:metabolism; D004441:Ecdysterone / Q000031:analogs & derivatives / Q000494:pharmacology; D005260:Female; D016193:G1 Phase / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3; D018969:Insulin-Like Growth Factor Binding Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
D050756:Cyclin-Dependent Kinase Inhibitor Proteins; C515497:IGFBP3 protein, human; D018972:Insulin-Like Growth Factor Binding Protein 3; D018969:Insulin-Like Growth Factor Binding Proteins; D012333:RNA, Messenger; D019938:Cyclin D1; D004441:Ecdysterone; C008999:ponasterone A; C495894:CDK2 protein, human; C495900:CDK4 protein, human; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4