title: Inhibition of Rac1 GTPase downregulates vascular endothelial growth factor receptor-2 expression by suppressing Sp1-dependent DNA binding in human endothelial cells.
abstract: RhoA, Rac1 and CDC42 are small GTP-binding proteins of the Rho family that play a crucial role in regulation of the actin-based cytoskeleton. In addition to cell growth regulation, they are implicated in transcriptional activation, oncogenic transformation and angiogenesis. The small Rho-GTPases have been linked to vascular endothelial growth factor (VEGF)-induced signalling pathways, but their role has not yet been elucidated. As signalling via the VEGF receptor-2 (VEGFR2) pathway is critical for angiogenic responses in cancer, wound repair and ischaemic and inflammatory diseases, we investigated whether the small Rho-GTPase Rac1 influences VEGFR2 expression in human endothelial cells. In this study, we show that a dominant negative Rac1 expression vector led to a pronounced decrease in VEGFR2 mRNA and protein expression. To identify minimal promoter requirements and potential applications of the small Rho-GTPases, we used VEGFR2 promoter-reporter gene constructs containing various deletions. The inhibitory effects of dominant negative Rac1 on the transcriptional activity of the VEGFR2 promoter localized to an element between -77 and -60 that contains an Sp1 transcription factor binding site. Electrophoretic mobility shift assays demonstrated that constitutive Sp1-dependent DNA binding decreased with Rac1 inhibition. Hence, repression of the small Rho GTPase Rac1 seems to be an additional critical molecular mechanism in the regulation of VEGFR2 expression.
authors.affiliation: Department of Dermatology, Klinikum der Johann Wolfgang Goethe-Universität, Frankfurt am Main, Germany. markus.meissner@kgu.de
authors.forename: Markus
authors.initials: M
authors.lastname: Meissner
authors.forename: Despina
authors.initials: D
authors.lastname: Michailidou
authors.forename: Monika
authors.initials: M
authors.lastname: Stein
authors.forename: Igor
authors.initials: I
authors.lastname: Hrgovic
authors.forename: Roland
authors.initials: R
authors.lastname: Kaufmann
authors.forename: Jens
authors.initials: J
authors.lastname: Gille
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D001665:Binding Sites / Q000235:genetics; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D042783:Endothelial Cells / Q000378:metabolism*; D015870:Gene Expression / Q000235:genetics; D006801:Humans; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000276:immunology / Q000378:metabolism*; D014162:Transfection; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; C107953:RAC1 protein, human; D034741:RNA, Small Interfering; D016329:Sp1 Transcription Factor; D004247:DNA; D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D020830:rac1 GTP-Binding Protein