NHERF links the N-cadherin/catenin complex to the platelet-derived growth factor receptor to modulate the actin cytoskeleton and regulate cell motility.
Using phage display, we identified Na+/H+ exchanger regulatory factor (NHERF)-2 as a novel binding partner for the cadherin-associated protein, beta-catenin. We showed that the second of two PSD-95/Dlg/ZO-1 (PDZ) domains of NHERF interacts with a PDZ-binding motif at the very carboxy terminus of beta-catenin. N-cadherin expression has been shown to induce motility in a number of cell types. The first PDZ domain of NHERF is known to bind platelet-derived growth factor-receptor beta (PDGF-Rbeta), and the interaction of PDGF-Rbeta with NHERF leads to enhanced cell spreading and motility. Here we show that beta-catenin and N-cadherin are in a complex with NHERF and PDGF-Rbeta at membrane ruffles in the highly invasive fibrosarcoma cell line HT1080. Using a stable short hairpin RNA system, we showed that HT1080 cells knocked down for either N-cadherin or NHERF had impaired ability to migrate into the wounded area in a scratch assay, similar to cells treated with a PDGF-R kinase inhibitor. Cells expressing a mutant NHERF that is unable to associate with beta-catenin had increased stress fibers, reduced lamellipodia, and impaired cell migration. Using HeLa cells, which express little to no PDGF-R, we introduced PDGF-Rbeta and showed that it coimmunoprecipitates with N-cadherin and that PDGF-dependent cell migration was reduced in these cells when we knocked-down expression of N-cadherin or NHERF. These studies implicate N-cadherin and beta-catenin in cell migration via PDGF-R-mediated signaling through the scaffolding molecule NHERF.
Theisen
Christopher S
CS
Department of Biochemistry and Molecular Biology, Eppley Cancer Center, University of Nebraska Medical Center, Omaha, NE 68198-7696, USA.
Wahl
James K
JK
Johnson
Keith R
KR
Wheelock
Margaret J
MJ
D000199:Actins / Q000378:metabolism* / Q000648:ultrastructure; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D015820:Cadherins / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement; D003599:Cytoskeleton / Q000378:metabolism; D005354:Fibrosarcoma / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000235:genetics / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
D000199:Actins; D015703:Antigens, CD; C050361:CDH2 protein, human; D015820:Cadherins; D046912:Multiprotein Complexes; D010750:Phosphoproteins; D017923:Sodium-Hydrogen Exchangers; D051176:beta Catenin; C093376:sodium-hydrogen exchanger regulatory factor; D020797:Receptor, Platelet-Derived Growth Factor beta