Insulin-like growth factor binding protein (IGFBP)-3 levels in conditioned media of Hs578T human breast cancer cells are post-transcriptionally regulated

Growth Regul. 1993 Mar;3(1):84-7.

Abstract

Hs578T human breast cancer cells secrete insulin-like growth factor binding protein (IGFBP)-3 (41-kDa and 39-kDa) and IGFBP-4 (24-kDa) as major BP species. In addition, cell surface-associated IGFBP-3 is demonstrable by use of cell monolayer affinity cross-linking or by employing immunoperoxidase staining of the cell surface with specific polyclonal anti-human IGFBP-3 antibodies (alpha IGFBP-3gl and alpha IGFBP-3ngl). In this study, we have demonstrated that regulation of Hs578T IGFBP-3 by IGF peptides is specific, non-receptor mediated, and post-translational by showing: 1) dose-dependent increase of IGFBP-3 in conditioned media(CM) following addition of IGF-I and IGF-II (maximum 8-13-fold increase at 100 ng/ml concentration), but not by insulin up to 1 microgram/ml; 2) confirmation of IGF-induced increases in CM concentrations of IGFBP-3 by means of Western ligand blot, affinity cross-linking, and IGFBP-3-specific radioimmunoassay; 3) increase of IGFBP-3 in CM by addition of IGF analogs which retain full affinity for IGFBPs ([Leu27]IGF-II and [Tyr55,Gln56]IGF-I), but not by IGF analogs which have significantly decreased affinity for IGFBPs ([Gln3,Ala4,Tyr15,Leu16]IGF-I, [Gln6,Ala7,Tyr18,Leu19,Leu27]IGF-II and IGF-I/insulin hybrid); 4) no change in IGFBP-3 mRNA following addition of IGFs; 5) existence of metal ion-dependent IGFBP-3 specific protease in CM and protection of IGFBP-3 from protease by formation of [IGF:IGFBP-3] complexes; and 6) release of cell surface-associated IGFBP-3 into CM by addition of IGF peptides. These studies demonstrate that IGF peptides regulate CM concentrations of IGFBP-3 through non-receptor mediated events, including dissociation of cell surface-associated IGFBP-3 and protection of IGFBP-3 from protease activity.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Blotting, Northern
  • Blotting, Western
  • Breast Neoplasms
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism*
  • Culture Media, Conditioned
  • Densitometry
  • Endopeptidases / metabolism
  • Humans
  • Insulin / physiology
  • Insulin-Like Growth Factor Binding Proteins
  • Protein Biosynthesis
  • Receptor, IGF Type 2 / metabolism
  • Somatomedins / metabolism*
  • Transcription, Genetic
  • Tumor Cells, Cultured

Substances

  • Carrier Proteins
  • Culture Media, Conditioned
  • Insulin
  • Insulin-Like Growth Factor Binding Proteins
  • Receptor, IGF Type 2
  • Somatomedins
  • Endopeptidases
  • insulin-like growth factor binding protein-3 protease