Tumor necrosis factor-alpha eliminates binding of NF-Y and an octamer-binding protein to the lipoprotein lipase promoter in 3T3-L1 adipocytes.
TNF alpha has been shown to reduce lipoprotein lipase (LPL) activity in adipose tissue. Regulation of LPL by TNF alpha occurs at the level of LPL gene transcription and posttranscriptionally. To elucidate further the transcriptional mechanism of TNF alpha inhibition of LPL gene transcription, transfection analysis was used to locate the site(s) of the LPL promoter that imparts the TNF alpha response. Transient transfections using LPL promoter deletions fused to luciferase in differentiated 3T3-L1 cells with and without TNF alpha treatment indicated that a DNA region downstream of -180 bp confers the TNF alpha effect. Electrophoretic mobility shift assays using two 32P-labeled LPL probes spanning the region between -180 and +44 bp revealed the loss of several LPL DNA-protein interactions after TNF alpha treatment, including the binding of NF-Y to the CCAAT box and a protein to the octamer consensus sequence. Protein binding to the OCT-1 consensus sequence is unaffected until after 4 h of TNF alpha treatment. In addition, the amount of mRNA for OCT-1 is not altered with TNF alpha treatment. These results indicate that TNF alpha regulates at least two DNA-binding proteins on the proximal promoter, thereby inhibiting LPL gene transcription.
Morin
C L
CL
Department of Endocrinology, Metabolism, and Diabetes, University of Colorado Health Sciences Center, Denver 80262, USA.
Schlaepfer
I R
IR
Eckel
R H
RH
D016475:3T3 Cells; D017667:Adipocytes / Q000187:drug effects*; D000818:Animals; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D051863:Host Cell Factor C1; D008071:Lipoprotein Lipase / Q000096:biosynthesis / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D050811:Octamer Transcription Factor-1; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; C497693:Hcfc1 protein, mouse; D051863:Host Cell Factor C1; D009687:Nuclear Proteins; D050811:Octamer Transcription Factor-1; C494166:Pou2f1 protein, mouse; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D008071:Lipoprotein Lipase