Fibronectin binds insulin-like growth factor-binding protein 5 and abolishes Its ligand-dependent action on cell migration.
Insulin-like growth factor-binding protein 5 (IGFBP-5) is a secreted protein that binds to insulin-like growth factors (IGFs) and modulates IGF actions on cell proliferation, differentiation, survival, and motility. IGFBP-5 also regulates these cellular events through IGF-independent mechanisms. To elucidate the molecular mechanisms governing these diverse actions of IGFBP-5, we screened a human cDNA library by a yeast two-hybrid system using IGFBP-5 as bait and identified fibronectin (FN) as a potential IGFBP-5-interacting partner. The complex formation of IGFBP-5 and FN was established by glutathione S-transferase pull-down, solution, and solid phase binding assays using glutathione S-transferase-IGFBP-5 and native IGFBP-5 in vitro and by co-immunoprecipitation in vivo. Binding assay using deletion mutants indicated that the IGFBP-5 C domain binds to the 10th and 11th type I repeats of FN. IGFBP-5 potentiated IGF-I-induced cell migration in FN-null, but not in wild-type, mouse embryonic cells. When FN was reintroduced either as an adhesive substrate or in solution to the FN-null cells, the potentiating effect of IGFBP-5 on IGF-I-induced cell migration was abolished. Binding of IGFBP-5 to FN had no effect on the ability of IGFBP-5 to bind IGF-I, but it increased the proteolytic degradation of IGFBP-5. Inhibition of IGFBP-5 proteolysis restored the potentiating effect of IGFBP-5. These results suggest that FN and IGFBP-5 bind to each other, and this binding negatively regulates the ligand-dependent action of IGFBP-5 by triggering IGFBP-5 proteolysis.
Xu
Qijin
Q
Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109-1048, USA.
Yan
Ben
B
Li
Shenghua
S
Duan
Cunming
C
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D005353:Fibronectins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D008024:Ligands; D051379:Mice; D018345:Mice, Knockout; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques
D018076:DNA, Complementary; D005353:Fibronectins; D018975:Insulin-Like Growth Factor Binding Protein 5; D008024:Ligands; D011993:Recombinant Fusion Proteins; D007334:Insulin-Like Growth Factor I