A non-IGF binding mutant of IGFBP-3 modulates cell function in breast epithelial cells

https://doi.org/10.1016/S0006-291X(02)00569-7Get rights and content

Abstract

We demonstrated previously that IGFBP-3 alone had no effect on cell death, but dramatically modulated apoptosis in Hs578T IGF non-responsive cells. We investigated whether a non-IGF binding mutant of IGFBP-3 retained its intrinsic actions in this cell line, prior to investigating its actions in IGF-responsive cells (MCF-7 and MCF-10A). In the Hs578T cells, the ceramide analogue, C2-induced apoptosis, non-glycosylated, glycosylated or mutant IGFBP-3 alone had no effect but on co-incubation with C2, all forms of IGFBP-3 markedly accentuated triggered apoptosis. In MCF-7 cells, IGFBP-3 was unable to modulate C2-induced death. In the MCF-10A cells, IGFBP-3 acted as a potent survival factor. IGFBP-3 also affected cell growth in the MCF-10A cells (inhibiting at low doses but increasing growth at higher concentrations). These actions of IGFBP-3 in the MCF-10A cells were independent of IGF-1. IGFBP-3 has differential IGF-independent effects on cell death and growth in normal breast and breast cancer cells.

Section snippets

Materials and methods

Materials. Insulin-like growth factor-I (IGF-I) was purchased from Gropep (Adelaide, Australia). An IGF-I receptor antagonist (IGFIR-AT) was purchased from Immunological and Biochemical Testsystems (IBT) GmbH (Reutlingen, Germany). The IGFIR-AT is a peptide that inhibits the autophosphorylation of the IGF-IR and has been shown to inhibit the proliferation of a number of cell lines in a dose-dependent manner [22]. Non-glycosylated (ng) IGFBP-3 was a gift from Dr. C.A. Macck (Celtrix

Effects of IGFBP-3 on C2-induced apoptosis in IGF-non-responsive Hs578T cells

In the Hs578T cell line, C2 induced a significant (P<0.05) increase in cell death from 10.5% to 23.8% whereas ng, gly, or mut IGFBP-3 alone had no effect in comparison to controls. As demonstrated previously [9], we confirmed that ng IGFBP-3 was able to significantly (P<0.05) accentuate C2-induced cell death from 23.8% to 47.3%. We now demonstrate further that gly and mut IGFBP-3 can similarly (P<0.05 for each) enhance C2-induced cell death from 23.8% to 44.1% and 40.2%, respectively (Fig. 1A).

Discussion

We have demonstrated in the Hs578T IGF non-responsive cell line that like ng IGFBP-3, gly, and mutant IGFBP-3 can also accentuate C2-induced cell death in an IGF-independent manner. Having established that the non-IGF binding mutant of IGFBP-3 maintained its intrinsic actions on cell death, we then examined the effects of this mutant in the IGF responsive MCF-7 and MCF10-A cell lines. We determined in the MCF-7 cell line that IGFBP-3 had no effect on C2-induced death, whereas in the MCF-10A

Acknowledgements

We wish to thank the Association for International Cancer Research and the Needham Cooper Trust for supporting this work.

References (39)

  • Y Imai et al.

    Substitutions for hydrophobic amino acids in the N-terminal domains of IGFBP-3 and -5 markedly reduce IGF-I binding and alter their biologic actions

    J. Biol. Chem.

    (2000)
  • Z.P Gill et al.

    Insulin-like growth factor-binding protein IGFBP-3 predisposes breast cancer cells to programmed cell death in a non-IGF-dependent manner

    J. Biol. Chem.

    (1997)
  • L.A Maile et al.

    Insulin-like growth factor binding protein (IGFBP) proteolysis: occurrence, identification, role and regulation

    Growth Horm. IGF Res.

    (1999)
  • P Angelloz-Nicoud et al.

    Prostate carcinoma (PC-3) cell proliferation is stimulated by the 22–25-kDa proteolytic fragment (1–160) and inhibited by the 16-kDa fragment (1–95) of recombinant human insulin-like growth factor binding protein-3

    Growth Horm. IGF Res.

    (1998)
  • S Fanayan et al.

    Growth inhibition by insulin-like growth factor-binding protein-3 in T47D breast cancer cells requires transforming growth factor-beta (TGF-beta) and the type II TGF-beta receptor

    J. Biol. Chem.

    (2000)
  • J.I Jones et al.

    Insulin-like growth factors and their binding proteins: biological actions

    Endocr. Rev.

    (1995)
  • R.C Baxter

    IGF binding protein-3 and the acid-labile subunit: formation of the ternary complex in vitro and in vivo

    Adv. Exp. Med. Biol.

    (1993)
  • W.F Blum et al.

    Insulin-like growth factor I (IGF-I)-binding protein complex is a better mitogen than free IGF-I

    Endocrinology

    (1989)
  • O Ritvos et al.

    Insulin-like growth factor (IGF) binding protein from human decidua inhibits the binding and biological action of IGF-I in cultured choriocarcinoma cells

    Endocrinology

    (1988)
  • Cited by (28)

    • IGFBP-3: A cell fate pivot in cancer and disease

      2014, Growth Hormone and IGF Research
      Citation Excerpt :

      This observation led to multifarious studies reporting IGF-independent apoptotic and growth-inhibitory effects of IGFBP-3. These have been shown through the addition or expression of non-IGF-binding IGFBP-3 analogues, culturing cells in serum/growth factor free conditions or through the use of non-IGF-responsive cells [56,106,115,119]. While there are a few reports of IGFBP-3 inducing apoptosis on its own, the majority of studies show IGFBP-3 acting as an induced molecule accentuating growth inhibitory and/or pro-apoptotic responses to extrinsic cues such as serum withdrawal, cytotoxic or cytostatic ligands or pharmacological induction of stress [35,114,136].

    • N-Acetylgalactosaminyltransferase 14, a novel insulin-like growth factor binding protein-3 binding partner

      2007, Biochemical and Biophysical Research Communications
      Citation Excerpt :

      Insulin-like growth factor binding protein (IGFBP)-3 has intrinsic anti-proliferative and pro-apoptotic functions that are independent of IGF binding. These effects are demonstrable with IGFBP-3 fragments that have the reduced IGF-I binding affinity and using cell lines that either lack IGF-I receptors or are unresponsive to IGF-I [11,12]. Cross-linking experiments indicate that IGFBP-3 is able to bind to a variety of cell surface proteins, including the type V transforming growth factor-β receptor [13].

    View all citing articles on Scopus
    View full text