THE HUMAN INSULIN-LIKE GROWTH-FACTOR (IGF) BINDING PROTEIN-3 INHIBITSTHE GROWTH OF FIBROBLASTS WITH A TARGETED DISRUPTION OF THE IGF-I RECEPTOR GENE

The insulin-like growth factors (IGFs) are important mitogens that exe rt their proliferative effects on cells via the type I IGF receptors ( IGF-R). The IGFs also associate with IGF binding proteins (IGFBPs). IG F-inhibitory, IGF-stimulatory, and IGF-independent effects of IGFBPs o n cell growth have been reported. We have asked whether the IGFBP-3 ha s an effect on cell growth, which is independent of its ability to bin d IGF-I and thus inhibit the activation of the IGF-I receptor. For thi s purpose, we have used a fibroblast cell line (R(-)cells) derived fro m mouse embryos homozygous for a targeted disruption of the IGF-R gene . When compared with wild type cells (W), which bind IGF-I with high a ffinity and specificity, R(-)cells transfected with a mammalian expres sion vector containing the human (h) IGFBP-3 cDNA were selected (R(-)/ BP3) and found to express hlGFBP-3 mRNA (detected by Northern blots) a nd to secrete hlGFBP-3 protein [detected by Western ligand blotting (W LB), immunoblotting, and immunoprecipitation as well as immunofluoresc ence confocal microscopy]. Growth of five different R(-)cells, and 10- fold slower compared with W cells, grown under identical conditions. C onfluent R(-)cells. These experiments show that the overexpression of IGFBP-3 has an inhibitory effect on cell growth which does not involve IGF binding to, or signal transduction via, the type I IGF-R. We conc lude that the cellular production of IGFBPs serves as a negative regul ator of cell proliferation which involves a cellular signaling pathway separate from the IGF-IGF-R system.