title: CD99 suppresses osteosarcoma cell migration through inhibition of ROCK2 activity.
abstract: CD99, a transmembrane protein encoded by MIC2 gene is involved in multiple cellular events including cell adhesion and migration, apoptosis, cell differentiation and regulation of protein trafficking either in physiological or pathological conditions. In osteosarcoma, CD99 is expressed at low levels and functions as a tumour suppressor. The full-length protein (CD99wt) and the short-form harbouring a deletion in the intracytoplasmic domain (CD99sh) have been associated with distinct functional outcomes with respect to tumour malignancy. In this study, we especially evaluated modulation of cell-cell contacts, reorganisation of the actin cytoskeleton and modulation of signalling pathways by comparing osteosarcoma cells characterised by different metastasis capabilities and CD99 expression, to identify molecular mechanisms responsible for metastasis. Our data indicate that forced expression of CD99wt induces recruitment of N-cadherin and β-catenin to adherens junctions. In addition, transfection of CD99wt inhibits the expression of several molecules crucial to the remodelling of the actin cytoskeleton, such as ACTR2, ARPC1A, Rho-associated, coiled-coil containing protein kinase 2 (ROCK2) as well as ezrin, an ezrin/radixin/moesin family member that has been clearly associated with tumour progression and metastatic spread in osteosarcoma. Functional studies point to ROCK2 as a crucial intracellular mediator regulating osteosarcoma migration. By maintaining c-Src in an inactive conformation, CD99wt inhibits ROCK2 signalling and this leads to ezrin decrease at cell membrane while N-cadherin and β-catenin translocate to the plasma membrane and function as main molecular bridges for actin cytoskeleton. Taken together, we propose that the re-expression of CD99wt, which is generally present in osteoblasts but lost in osteosarcoma, through inhibition of c-Src and ROCK2 activity, manages to increase contact strength and reactivate stop-migration signals that counteract the otherwise dominant promigratory action of ezrin in osteosarcoma cells.
authors: Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.
authors: C
authors: C
authors: Zucchini
authors: 1] CRS Development of Biomolecular Therapies, Bologna, Italy [2] Experimental Oncology Laboratory, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: M C
authors: MC
authors: Manara
authors: 1] CRS Development of Biomolecular Therapies, Bologna, Italy [2] Experimental Oncology Laboratory, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: R S
authors: RS
authors: Pinca
authors: Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.
authors: P
authors: P
authors: De Sanctis
authors: 1] Experimental Oncology Laboratory, Istituto Ortopedico Rizzoli, Bologna, Italy [2] PROMETEO Laboratory, STB, RIT Department, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: C
authors: C
authors: Guerzoni
authors: CRS Development of Biomolecular Therapies, Bologna, Italy.
authors: M
authors: M
authors: Sciandra
authors: Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.
authors: P-L
authors: PL
authors: Lollini
authors: Department of Biomedical and Neuromotor Sciences, University of Bologna, Bologna, Italy.
authors: G
authors: G
authors: Cenacchi
authors: Experimental Oncology Laboratory, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: P
authors: P
authors: Picci
authors: Department of Experimental, Diagnostic and Specialty Medicine, University of Bologna, Bologna, Italy.
authors: L
authors: L
authors: Valvassori
authors: 1] CRS Development of Biomolecular Therapies, Bologna, Italy [2] Experimental Oncology Laboratory, Istituto Ortopedico Rizzoli, Bologna, Italy [3] PROMETEO Laboratory, STB, RIT Department, Istituto Ortopedico Rizzoli, Bologna, Italy.
authors: K
authors: K
authors: Scotlandi
mesh_terms: D000071443:12E7 Antigen; D008841:Actin Cytoskeleton / Q000378:metabolism; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002448:Cell Adhesion / Q000235:genetics; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D003598:Cytoskeletal Proteins; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D046529:Microscopy, Electron, Transmission; D009361:Neoplasm Invasiveness / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D012516:Osteosarcoma / Q000235:genetics* / Q000378:metabolism* / Q000473:pathology; D034741:RNA, Small Interfering; D060888:Real-Time Polymerase Chain Reaction; D014162:Transfection; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism*; D000099149:Ezrin
chemical_list: D000071443:12E7 Antigen; D015703:Antigens, CD; D015820:Cadherins; D015815:Cell Adhesion Molecules; D003598:Cytoskeletal Proteins; D034741:RNA, Small Interfering; D054460:rho-Associated Kinases; D000099149:Ezrin; C061828:CD99 protein, human; C515852:ROCK2 protein, human