Surface-bound plasmin induces selective proteolysis of insulin-like-growth-factor (IGF)-binding protein-4 (IGFBP-4) and promotes autocrine IGF-II bio-availability in human colon-carcinoma cells.
Limited proteolysis of insulin-like-growth-factor (IGF)-binding proteins (IGFBPs) represents a key process to modulate IGF bio-availability at the cellular level. In human colon carcinomas, urokinase-type plasminogen activator (u-PA) produced by stroma cells can bind to cancer-cell-associated u-PA receptor (u-PAR), and then catalyze the conversion of plasminogen (Pg) into plasmin (Pm). We therefore investigated the interplay between the IGF and Pm systems in the HT29-D4 human colon-carcinoma-cell model. HT29-D4 cells secreted IGF-II totally complexed to IGFBP-2, IGFBP-4 and IGFBP-6. Approximately 15% of IGFBP-4 was associated with the extracellular matrix. HT29-D4 cells produced neither u-PA- nor IGFBP-specific proteases. However, activation of Pm at the HT29-D4 cell surface obtained by the sequential addition of exogenous u-PA and Pg to mimic the stromal complementation induced selective proteolysis targeted to IGFBP-4 only (>95%). IGFBP-2 and IGFBP-6, though sensitive to proteolysis by soluble Pm, were not altered by cell-bound Pm. IGFBP-4 proteolysis yielded 18- and 14-kDa immunoreactive fragments which were not detectable by Western ligand blotting, indicating that they bound IGF-II with poor affinity. Release of IGF-II from IGF-II-IGFBP complexes after IGFBP-4 proteolysis by cell-bound Pm was indicated by the observation that approximately 20% of the 125I-IGF-II initially associated with endogenous IGFBP in reconstituted complexes was transferred to HT29-D4 cell-surface IGF-I receptors. These results suggest that IGFBP-4 proteolysis by cell-bound Pm can promote autocrine/paracrine IGF-II bio-availability in colon-cancer cells. This may have important consequences on the behavior of cancer cells at the interface between stroma and malignant cells in carcinomas of the colon in vivo.
Remacle-Bonnet
M M
MM
Unite Interactions entre Systemes Proteiques et Differenciation dans la Cellule Tumorale, Faculte de Medecine, URA CNRS 1924, Marseille, France.
Garrouste
F L
FL
Pommier
G J
GJ
D000230:Adenocarcinoma / Q000378:metabolism*; D007611:Aprotinin / Q000494:pharmacology; D015153:Blotting, Western; D003110:Colonic Neoplasms / Q000378:metabolism*; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism; D010450:Endopeptidases / Q000032:analysis / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism*; D006801:Humans; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000032:analysis / Q000378:metabolism*; D007335:Insulin-Like Growth Factor II / Q000032:analysis / Q000378:metabolism*; D010958:Plasminogen / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D017077:Culture Media, Conditioned; D018974:Insulin-Like Growth Factor Binding Protein 4; D007335:Insulin-Like Growth Factor II; D010958:Plasminogen; D007611:Aprotinin; D010450:Endopeptidases; D005341:Fibrinolysin