Insulin-like growth factor-I (IGF-I) regulates IGFBP-3 and IGFBP-4 by multiple mechanisms in A549 human adenocarcinoma cells.

Abstract

The insulin-like growth factors (IGF-I and IGF-II) participate in the control of cell proliferation in normal and neoplastic lung cells. To examine the role of IGF binding proteins (IGFBPs) in modulating IGF actions in lung, we examined the production and regulation of IGFBPs from A549 cells, a human adenocarcinoma-derived lung cell line. Ligand blot and immunoblot analysis of conditioned media (CM) from A549 cells demonstrated IGFBP bands of relative molecular mass (M(r)) approximately 39-43,000 (IGFBP-3), 34,000 (IGFBP-2), 30,000 (IGFBP-1), and 24,000 (IGFBP-4). IGFBP-3 abundance in A549 cell CM increased following exposure to IGF-I and IGF-II (3.0- and 1.8-fold, respectively) without a change in IGFBP-3 transcript abundance, suggesting IGFBP-3 is post-transcriptionally regulated. Cycloheximide almost completely abrogated the IGF-I-stimulated increase in CM IGFBP-3, suggesting that ongoing protein synthesis is necessary for the IGF-I-stimulated increase in IGFBP-3 abundance. Increases in IGFBP-3 occurred by at least two mechanisms, through activation of the type 1 IGF receptor and by a type 1 IGF receptor independent mechanism. The increase in IGFBP-3 was due, in part, to activation of the type 1 IGF receptor because blocking type 1 IGF receptor activation with an antibody (alpha IR3) diminished the IGF-I-induced increase in IGFBP-3 and insulin, at doses that stimulate the type 1 IGF receptor, increased IGFBP-3 abundance. The increase in IGFBP-3 was partially independent of type 1 IGF receptor activation because [QAYL]-IGF-I, an analog of IGF-I that binds the type 1 IGF receptor but not IGFBP-3, was less potent than IGF-I in stimulating IGFBP-3 abundance, and IGF-II, which binds IGFBP-3 normally, but binds the type 1 IGF receptor with lower affinity than IGF-I, was nearly equipotent to IGF-I in its stimulation of IGFBP-3 accumulation at low concentrations. These results suggest that ligand binding decreases IGFBP-3 clearance or increases IGFBP-3 accumulation in CM. IGF-I decreased IGFBP-4 abundance in A549 cell CM without decreasing IGFBP-4 mRNA transcripts and without increasing the amount of cell-associated IGFBP-4. To determine whether the decrease in IGFBP-4 was due to increased degradation, cell-free CM was incubated with and without IGF-I, and IGFBP-4 abundance measured by ligand and immunoblot analyses.(ABSTRACT TRUNCATED AT 400 WORDS)