v-Jun downregulates the SPARC target gene by binding to the proximal promoter indirectly through Sp1 3

• Published in: Oncogene Vol. 22; no. 26; pp. 4047 - 4061
• Main Authors: CHAMBOREDON, Sandrine, BRIGGS, Joseph, VIAL, Emmanuel, HURAULT, Julien, GALVAGNI, Federico, OLIVIERO, Salvatore, BOS, Timothy, CASTELLAZZI, Marc
• Format: Journal Article
• Language: English
• Published: Basingstoke Nature Publishing 26.06.2003; Nature Publishing Group
• Subjects: Amino Acid Sequence; Animals; Biological and medical sciences; Blotting, Western; Cell Line; Cell Nucleus - metabolism; Cells, Cultured; Chick Embryo; Chromatin; Chromatin - metabolism; Deoxyribonucleic acid; Dimerization; DNA; DNA - metabolism; DNA, Complementary - metabolism; DNA-Binding Proteins - metabolism; Dose-Response Relationship, Drug; Down-Regulation; Drosophila; Electrophoretic mobility; Embryo fibroblasts; Extracellular matrix; Fibroblasts; Fibroblasts - metabolism; Fundamental and applied biological sciences. Psychology; Genes; Glutathione Transferase - metabolism; Immunoprecipitation; Laboratories; Luciferases - metabolism; Matrix protein; Models, Biological; Molecular and cellular biology; Molecular Sequence Data; Oncogene Protein p65(gag-jun) - metabolism; Osteonectin; Osteonectin - metabolism; Plasmids - metabolism; Precipitin Tests; Promoter Regions, Genetic; Protein Binding; Proteins; RNA, Messenger - metabolism; Sequence Homology, Amino Acid; Sp1 protein; Sp1 Transcription Factor - metabolism; Sp3 Transcription Factor; Transcription activation; Transcription factors; Transcription Factors - metabolism; Transcription, Genetic; Transcriptional Activation; Transfection; Tumorigenesis; Tumors; V-Jun protein; cell transformation; SPARC; Sp1; Sp3; v-Jun; Genetics; primary chick embryo fibroblasts
• ISSN: 0950-9232; 1476-5594
• DOI: 10.1038/sj.onc.1206713

Transformation of chick embryo fibroblasts by the v-Jun oncoprotein correlates with a downregulation of the extracellular matrix protein SPARC and repression of the corresponding mRNA. Repression of SPARC contributes to the oncogenic process by facilitating tumor development in vivo. A proximal promoter fragment, designated -124/+16, is responsible for high constitutive activity of the SPARC gene and is the target of repression by v-Jun. In this paper, using electrophoretic mobility shift and pull-down assays in vitro, and transient transfections and chromatin immunoprecipitation assays in Sp1/3-deficient Drosophila SL2 cells and in chick embryo fibroblasts, we show that (i) Sp1 and/or Sp3 is required for constitutive activation of SPARC transcription, by binding directly to the GGA-rich -92/-57 fragment; and (ii) v-Jun does not bind -124/+16 directly, but binds to the GGA-rich fragment indirectly, most likely through a physical interaction with Sp1/3. Moreover, a transactivation-proficient v-Jun derivative, designated v-Jun/cebp/glz, which cannot bind Jun DNA motifs anymore and cannot heterodimerize, is still capable of downregulating SPARC efficiently. Taken together, these data strongly suggest that v-Jun downregulates SPARC through the formation of a DNA-Sp1/3-v-Jun, chromatin-associated complex.