Expression of dominant-negative form of Ets-1 suppresses fibronectin-stimulated cell adhesion and migration through down-regulation of integrin alpha5 expression in U251 glioma cell line.
Ets transcription factors are associated with tumor malignancy. We reported previously that the stable transfection of the dominant-negative form of Ets-1 (Ets-DN) in the glioma cell line U251 induced down-regulation of urokinase-type plasminogen activator mRNA expression and invasiveness (M. Nakada et al., J. Neuropathol. Exp. Neurol., 58: 329-334, 1999). Here we analyzed effects of Ets-DN expression on cell adhesion, migration, and phosphorylation of focal adhesion kinase. U251 cells expressing Ets-DN (U251-DN) showed reduced cell adhesion, spreading, and extension of actin stress fibers on dishes coated with fibronectin but not on dishes coated with collagen. Migration of U251-DN cells was found to be significantly inhibited compared with that of parental cells when examined by wound-induced migration assay on fibronectin-coated dishes. Phosphorylation levels of focal adhesion kinase in U251-DN cells were also attenuated on dishes coated with fibronectin. Reduced expression level of integrin alpha5 subunit in U251-DN cells was demonstrated by semiquantitative reverse transcription-PCR analysis. Semiquantitative reverse transcription-PCR of surgical samples of brain tumors revealed that the expression level of Ets-1 mRNA correlated with that of integrin alpha5 mRNA in glioma. The experimental metastatic ability of U251-DN cells examined in chick embryo was considerably lower than that of parental cells. These results suggest that Ets-1 contributes to glioma malignancy by up- regulating expression of the integrin alpha5 subunit, which composes integrin alpha5beta1 and mediates intracellular signaling and the subsequent acceleration of the invasive process, including cell adhesion and migration.
Kita
D
D
Department of Molecular Virology and Oncology, Kanazawa University, Kanazawa, Ishikawa 920-0934, Japan.
Takino
T
T
Nakada
M
M
Takahashi
T
T
Yamashita
J
J
Sato
H
H
D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis* / Q000235:genetics; D002448:Cell Adhesion; D002465:Cell Movement; D002642:Chick Embryo; D024042:Collagen Type I / Q000378:metabolism / Q000494:pharmacology; D015536:Down-Regulation; D005353:Fibronectins / Q000378:metabolism / Q000494:pharmacology*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015972:Gene Expression Regulation, Neoplastic; D005910:Glioma / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D005914:Globins / Q000235:genetics; D006801:Humans; D039482:Integrin alpha5; D039661:Integrin beta3; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism / Q000556:secondary; D010766:Phosphorylation; D010980:Platelet Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D050783:Proto-Oncogene Proteins c-ets; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D014162:Transfection; D014407:Tumor Cells, Cultured
D015703:Antigens, CD; D024042:Collagen Type I; C494041:ETS1 protein, human; D005353:Fibronectins; D039482:Integrin alpha5; D039661:Integrin beta3; D010980:Platelet Membrane Glycoproteins; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D012333:RNA, Messenger; D014157:Transcription Factors; D005914:Globins; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human