Interactions of cathepsin-D and insulin-like growth factor-II (IGF-II) on the IGF-II/mannose-6-phosphate receptor in human breast cancer cells and possible consequences on mitogenic activity of IGF-II

Mol Endocrinol. 1990 Sep;4(9):1327-35. doi: 10.1210/mend-4-9-1327.

Abstract

The lysosomal enzyme cathepsin-D (cath-D) and insulin-like growth factor-II (IGF-II), which share a common IGF-II/mannose-6-phosphate (M6P) transmembrane receptor, are both synthesized and secreted by breast cancer cells, upon which they might exert an intracrine/autocrine control on proliferation. We have evaluated the binding of 125I-immunopurified human cath-D in different breast cell membrane preparations. The concentration of high affinity M6P reversible binding sites (mean Kd, 0.85 nM) varied among the different breast cancer cells (0-0.82 pmol/mg membrane protein), but there was no correlation between the presence of steroid receptor and M6P-dependent binding. Cross-linking experiments with [125I]cath-D and [125I]IGF-II showed the formation of complexes with the 270,000 mol wt IGF-II/M6P receptor molecule which migrated, respectively, at 330,000 and 270,000 mol wt in 3-10% gradient sodium dodecyl sulfate-polyacrylamide gels. [125I]IGF-II cross-linking was increased by M6P (20% above control), whereas cath-D strongly inhibited IGF-II interaction by 80%. Conversely, IGF-II reduced [125I]cath-D cross-linking by 55%. Direct ligand binding on receptors transferred onto nitrocellulose sheets by Western blotting confirmed the interaction of both ligands on the same receptor molecule. By studying IGF-II's growth-promoting activity in these cells in a wide range of concentrations, we show that IGF-II triggers its mitogenic response via IGF-II/M6P receptor at low concentrations, whereas it is mainly acting via IGF-I receptor at high concentrations. Three lines of evidences lead us to that conclusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding, Competitive
  • Blotting, Western
  • Breast Neoplasms / metabolism*
  • Breast Neoplasms / pathology
  • Cathepsin D / metabolism*
  • Cathepsin D / pharmacology
  • Cell Division / drug effects
  • Cell Membrane / metabolism
  • Cross-Linking Reagents
  • Electrophoresis, Polyacrylamide Gel
  • Humans
  • Hydrogen-Ion Concentration
  • Insulin-Like Growth Factor II / metabolism*
  • Insulin-Like Growth Factor II / pharmacology
  • Mannosephosphates / metabolism
  • Molecular Weight
  • Receptor, IGF Type 2
  • Receptors, Cell Surface / metabolism*
  • Receptors, Somatomedin
  • Succinimides
  • Tumor Cells, Cultured

Substances

  • Cross-Linking Reagents
  • Mannosephosphates
  • Receptor, IGF Type 2
  • Receptors, Cell Surface
  • Receptors, Somatomedin
  • Succinimides
  • mannose-6-phosphate
  • Insulin-Like Growth Factor II
  • Cathepsin D
  • disuccinimidyl suberate