Insulin-like growth factor I controls a mutually exclusive association of RACK1 with protein phosphatase 2A and beta1 integrin to promote cell migration.
The WD repeat scaffolding protein RACK1 can mediate integration of the insulin-like growth factor I receptor (IGF-IR) and integrin signaling in transformed cells. To address the mechanism of RACK1 function, we searched for regulatory proteins that associate with RACK1 in an IGF-I-dependent manner. The serine threonine phosphatase protein phosphatase 2A (PP2A) was found associated with RACK1 in serum-starved cells, and it dissociated immediately upon stimulation with IGF-I. This dissociation of PP2A from RACK1 and an IGF-I-mediated decrease in cellular PP2A activity did not occur in cells expressing either the serine 1248 or tyrosine 1250/1251 mutants of the IGF-IR that do not interact with RACK1. Recombinant RACK1 could bind to PP2A in vitro and restore phosphatase activity to PP2A from IGF-I-stimulated cells. Ligation of integrins with fibronectin or Matrigel was sufficient to facilitate IGF-I-mediated dissociation of PP2A from RACK1 and also to recruit beta1 integrin as PP2A dissociated. By using TAT-fused N-terminal and C-terminal deletion mutants of RACK1, we determined that both PP2A and beta1 integrin interact in the C terminus of RACK1 within WD repeats 4 to 7. This suggests that integrin ligation displaces PP2A from RACK1. MCF-7 cells overexpressing RACK1 exhibited enhanced motility, which could be reversed by the PP2A inhibitor okadaic acid. Small interfering RNA-mediated suppression of RACK1 also decreased the migratory capacity of DU145 cells. Taken together, our findings indicate that RACK1 enhances IGF-I-mediated cell migration through its ability to exclusively associate with either beta1 integrin or PP2A in a complex at the IGF-IR.
Kiely
Patrick A
PA
Cell Biology Laboratory, Department of Biochemistry, BioSciences Institute, National University of Ireland, Cork, Ireland.
O'Gorman
Denise
D
Luong
Ken
K
Ron
Dorit
D
O'Connor
Rosemary
R
D000818:Animals; D002465:Cell Movement / Q000187:drug effects*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D015870:Gene Expression; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D019012:Integrin beta1 / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D008954:Models, Biological; D009363:Neoplasm Proteins / Q000378:metabolism*; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D054648:Protein Phosphatase 2; D034741:RNA, Small Interfering / Q000235:genetics; D017526:Receptor, IGF Type 1 / Q000172:deficiency; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D019012:Integrin beta1; D009363:Neoplasm Proteins; C419368:RACK1 protein, human; D034741:RNA, Small Interfering; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface; D019319:Okadaic Acid; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2; D019204:GTP-Binding Proteins