Heping Cao1, Joseph F Urban, Richard A Anderson. 1. Diet, Genomics, and Immunology Laboratory, Beltsville Human Nutrition Research Center, Agricultural Research Service, US Department of Agriculture, Beltsville, Maryland, USA. Heping.Cao@ars.usda.gov
Abstract
OBJECTIVES: Tristetraprolin (TTP) family proteins (TTP/ZFP36; ZFP36L1, ZFP36L2, ZFP36L3) destabilize adenylate uridylate-rich element-containing mRNAs encoding cytokines, such as tumor necrosis factor (TNF) and vascular endothelial growth factor (VEGF). Little is known about the expression and insulin regulation of TTP and related genes in adipocytes. We analyzed the relative abundance of TTP family mRNAs in 3T3-L1 adipocytes compared to RAW264.7 macrophages and investigated insulin effects on the expression of 43 genes in 3T3-L1 adipocytes. METHODS AND PROCEDURES: Insulin was added to mouse 3T3-L1 adipocytes. Relative abundance of mRNA levels was determined by quantitative real-time PCR. TTP and ZFP36L1 proteins were detected by immunoblotting. RESULTS: Zfp36l1 and Zfp36l2 genes were expressed at eight- to tenfold higher than Ttp in adipocytes. Zfp36l3 mRNA was detected at approximately 1% of Ttp mRNA levels in adipocytes and its low level expression was confirmed in RAW cells. Insulin at 10 and 100 nmol/l increased Ttp mRNA levels by five- to sevenfold, but decreased those of Zfp36l3 by 40% in adipocytes after a 30-min treatment. Immunoblotting showed that insulin induced TTP but did not affect ZFP36L1 protein levels in adipocytes. Insulin decreased mRNA levels of Vegf and a number of other genes in adipocytes. DISCUSSION: Insulin induced Ttp mRNA and protein expression and decreased Vegf mRNA levels in adipocytes. Zfp36l3 mRNA was detected, for the first time, in cells other than mouse placenta and extraembryonic tissues. This study established a basis for the investigation of TTP and VEGF genes in the regulation of obesity and suggested that Vegf mRNA may be a target of TTP in fat cells.
OBJECTIVES:Tristetraprolin (TTP) family proteins (TTP/ZFP36; ZFP36L1, ZFP36L2, ZFP36L3) destabilize adenylate uridylate-rich element-containing mRNAs encoding cytokines, such as tumor necrosis factor (TNF) and vascular endothelial growth factor (VEGF). Little is known about the expression and insulin regulation of TTP and related genes in adipocytes. We analyzed the relative abundance of TTP family mRNAs in 3T3-L1 adipocytes compared to RAW264.7 macrophages and investigated insulin effects on the expression of 43 genes in 3T3-L1 adipocytes. METHODS AND PROCEDURES: Insulin was added to mouse 3T3-L1 adipocytes. Relative abundance of mRNA levels was determined by quantitative real-time PCR. TTP and ZFP36L1 proteins were detected by immunoblotting. RESULTS:Zfp36l1 and Zfp36l2 genes were expressed at eight- to tenfold higher than Ttp in adipocytes. Zfp36l3 mRNA was detected at approximately 1% of Ttp mRNA levels in adipocytes and its low level expression was confirmed in RAW cells. Insulin at 10 and 100 nmol/l increased Ttp mRNA levels by five- to sevenfold, but decreased those of Zfp36l3 by 40% in adipocytes after a 30-min treatment. Immunoblotting showed that insulin induced TTP but did not affect ZFP36L1 protein levels in adipocytes. Insulin decreased mRNA levels of Vegf and a number of other genes in adipocytes. DISCUSSION: Insulin induced Ttp mRNA and protein expression and decreased Vegf mRNA levels in adipocytes. Zfp36l3 mRNA was detected, for the first time, in cells other than mouse placenta and extraembryonic tissues. This study established a basis for the investigation of TTP and VEGF genes in the regulation of obesity and suggested that Vegf mRNA may be a target of TTP in fat cells.