Matrix metalloproteinase-9 silencing by RNA interference triggers the migratory-adhesive switch in Ewing's sarcoma cells.
Enhanced expression of (pro)matrix metalloproteinase-9 (MMP-9) is associated with human tumor invasion and/or metastasis. COH cells derived from a highly invasive and metastatic Ewing's sarcoma constitutively express proMMP-9. Transfection of a double stranded RNA that targets the MMP-9 mRNA into COH cells depleted the corresponding mRNA and protein as demonstrated by reverse transcriptase-PCR, enzyme-linked immunosorbent assay, and gelatin zymography. proMMP-9 extinction resulted in the following: (i) decreased spreading on extracellular matrix (fibronectin, laminin, collagen IV)-coated surfaces, (ii) inhibition of migration toward fibronectin, and (iii) induced aggregation, which was specifically disrupted by a function-blocking E-cadherin antibody. MMP-9 knockdown concomitantly resulted in increased levels of surface E-cadherin, redistribution at the plasma membrane of beta-catenin, and its physical association with E-cadherin. Moreover, induction of E-cadherin-mediated adhesion was associated with RhoA activation and changes in paxillin cytoskeleton. Finally, an inhibitor of gelatinolytic activity of pro-MMP9 did not reduce COH cell migration confirming that the enzymatic property of COH MMP-9 was not required for migration toward fibronectin. Overall, our observations define a novel critical role for proMMP-9 in providing a cellular switch between stationary and migratory cell phases.
Sancéau
Josiane
J
Unité 365 INSERM, Institut Curie, 75248 Paris cedex 05, France.
Truchet
Sandrine
S
Bauvois
Brigitte
B
D015820:Cadherins / Q000378:metabolism; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D002465:Cell Movement; D003094:Collagen / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D005434:Flow Cytometry; D020868:Gene Silencing; D016158:Genes, p53; D006801:Humans; D007797:Laminin / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D011485:Protein Binding; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D012512:Sarcoma, Ewing / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D051176:beta Catenin
C495270:CTNNB1 protein, human; D015820:Cadherins; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D005353:Fibronectins; D007797:Laminin; D012333:RNA, Messenger; D015534:Trans-Activators; D051176:beta Catenin; D003094:Collagen; D020780:Matrix Metalloproteinase 9