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PMID: 9271438 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

The von Hippel-Lindau tumor suppressor gene product interacts with Sp1 to repress vascular endothelial growth factor promoter activity.

Molecular and cellular biology ·Vol. 17 ·No. 9 ·1997-09-00 ·Pages 5629-39

Mukhopadhyay D, Knebelmann B, Cohen HT, Ananth S, Sukhatme VP

Abstract

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/genetics,metabolism Enhancer Elements, Genetic Genes, Tumor Suppressor Humans Ligases Lymphokines/genetics,metabolism Promoter Regions, Genetic Proteins/metabolism RNA, Messenger/metabolism Sp1 Transcription Factor/metabolism 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
Chemicals
Endothelial Growth Factors Lymphokines Proteins RNA, Messenger Sp1 Transcription Factor Tumor Suppressor Proteins Vascular Endothelial Growth Factor A Vascular Endothelial Growth Factors Ubiquitin-Protein Ligases Von Hippel-Lindau Tumor Suppressor Protein Ligases VHL protein, human
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Mukhopadhyay D
Department of Medicine, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, Massachusetts 02215, USA.
Knebelmann B
Cohen H T
Ananth S
Sukhatme V P
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.17.9.5629
Published
1997-09-00
Pages
5629-39
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC232411
Subset
IM
Grants
NIDDK NIH HHS · DK02280 · United States
NIDDK NIH HHS · DK44921 · United States
External Links
PubMed source DOI: 10.1128/MCB.17.9.5629 PMC Full Text View journal details
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