The von Hippel-Lindau tumor suppressor gene product interacts with Sp1 to repress vascular endothelial growth factor promoter activity.
The von Hippel-Lindau tumor suppressor gene (VHL) has a critical role in the pathogenesis of clear-cell renal cell carcinoma (RCC), as VHL mutations have been found in both von Hippel-Lindau disease-associated and sporadic RCCs. Recent studies suggest that vascular endothelial growth factor (VEGF) mRNA is upregulated in RCC- and von ... Hippel-Lindau disease-associated tumors. We have therefore assessed the effect of the VHL gene product on VEGF expression. VEGF promoter-luciferase constructs were transiently cotransfected with a wild-type VHL (wt-VHL) vector in several cell lines, including 293 embryonic kidney and RCC cell lines. wt-VHL protein inhibited VEGF promoter activity in a dose-dependent manner up to 5- to 10-fold. Deletion analysis defined a 144-bp region of the VEGF promoter necessary for VHL repression. This VHL-responsive element is GC rich and specifically binds the transcription factor Sp1 in crude nuclear extracts. In Drosophila cells, cotransfected VHL represses Sp1-mediated activation but not basal activity of the VEGF promoter. We next demonstrated in coimmunoprecipitates that VHL and Sp1 were part of the same complex and, by using a glutathione-S-transferase-VHL fusion protein and purified Sp1, that VHL and Sp1 directly interact. Furthermore, endogenous VEGF mRNA levels were suppressed in permanent RCC cell lines expressing wt-VHL, and nuclear run-on studies indicated that VHL regulation of VEGF occurs at least partly at the transcriptional level. These observations support a new mechanism for VHL-mediated transcriptional repression via a direct inhibitory action on Sp1 and suggest that loss of Sp1 inhibition may be important in the pathogenesis of von Hippel-Lindau disease and RCC.
Mesh Terms:
Animals, COS Cells, Endothelial Growth Factors, Enhancer Elements, Genetic, Genes, Tumor Suppressor, Humans, Ligases, Lymphokines, Promoter Regions, Genetic, Proteins, RNA, Messenger, Sp1 Transcription Factor, Transcription, Genetic, Transfection, Tumor Cells, Cultured, Tumor Suppressor Proteins, Ubiquitin-Protein Ligases, Vascular Endothelial Growth Factor A, Vascular Endothelial Growth Factors, Von Hippel-Lindau Tumor Suppressor Protein
Animals, COS Cells, Endothelial Growth Factors, Enhancer Elements, Genetic, Genes, Tumor Suppressor, Humans, Ligases, Lymphokines, Promoter Regions, Genetic, Proteins, RNA, Messenger, Sp1 Transcription Factor, Transcription, Genetic, Transfection, Tumor Cells, Cultured, Tumor Suppressor Proteins, Ubiquitin-Protein Ligases, Vascular Endothelial Growth Factor A, Vascular Endothelial Growth Factors, Von Hippel-Lindau Tumor Suppressor Protein
Mol. Cell. Biol.
Date: Sep. 01, 1997
PubMed ID: 9271438
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