Insulin-like growth factor binding protein-2 mediates the inhibition of DNA synthesis by transforming growth factor-beta in mink lung epithelial cells.
Insulin-like growth factor binding protein-3 (IGFBP-3) has been proposed to mediate the growth inhibitory effects of transforming growth factor (TGF)-beta in breast and prostate cancer cells. Both TGF-beta and exogenous IGFBP-3 inhibit DNA synthesis in Mv1 mink lung epithelial cells (CCL64). The present study asks whether IGFBPs synthesized by CCL64 cells mediate growth inhibition by TGF-beta. CCL64 cells synthesize and secrete a single 34-kDa IGFBP that was identified as IGFBP-2 by immunoprecipitation and immunodepletion. Recombinant bovine IGFBP-2 inhibited CCL64 DNA synthesis in serum-free media in an IGF-independent manner. Coincubation with Leu(60)-IGF-I, an IGF-I analog that binds to IGFBPs with higher affinity than to IGF-I receptors, decreased the inhibition by bIGFBP-2. Leu(60)-IGF-I also decreased the inhibition of CCL64 DNA synthesis by TGF-beta by up to 70%, whereas Long-R3-IGF-I, an IGF-I analog with higher affinity for IGF-I receptors than for IGFBPs, did not decrease inhibition, suggesting that the effect of Leu(60)-IGF-I resulted from its forming complexes with endogenous IGFBPs. Leu(60)-IGF-I did not decrease TGF-beta stimulation of a Smad3-dependent reporter gene. Following incubation of intact CCL64 cells with bIGFBP-2 at 0 degrees C, bIGFBP-2 was recovered in membrane fractions; membrane association was abolished by coincubation with Leu(60)-IGF-I. If exogenous and secreted IGFBP-2 must bind to CCL64 cells to inhibit DNA synthesis, Leu(60)-IGF-I might reduce the inhibition of DNA synthesis by bIGFBP-2 or TGF-beta by inhibiting the association of IGFBP-2 in the media with CCL64 cells. Since TGF-beta does not increase IGFBP-2 abundance, we propose that TGF-beta sensitizes CCL64 cells to the latent growth inhibitory activity of endogenous IGFBP-2 by potentiating an intracellular IGFBP-2 signaling pathway or by promoting the association of secreted IGFBP-2 with the plasma membrane.
Dong
Feng
F
Growth and Development Section, Clinical Endocrinology Branch, National Institute of Diabetes and Digestive and Kidney Diseases/NIH, Bldg. 10, Room 8D12, Bethesda, MD 20892, USA.
Wu
Hai-Bin
HB
Hong
Jiang
J
Rechler
Matthew M
MM
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002417:Cattle; D002460:Cell Line; D006224:Cricetinae; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis*; D017930:Genes, Reporter; D006801:Humans; D018971:Insulin-Like Growth Factor Binding Protein 2 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D008168:Lung / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008907:Mink; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D020545:Respiratory Mucosa / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000494:pharmacology*
D017077:Culture Media, Conditioned; D018971:Insulin-Like Growth Factor Binding Protein 2; D019384:Nucleic Acid Synthesis Inhibitors; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D007334:Insulin-Like Growth Factor I; D004247:DNA