Leptin increases tissue inhibitor of metalloproteinase I (TIMP-1) gene expression by a specificity protein 1/signal transducer and activator of transcription 3 mechanism.
Leptin has properties of a profibrogenic cytokine. In liver, the activated hepatic stellate cell (HSC) is responsible for a net production of extracellular matrix. A key molecule synthesized is the tissue inhibitor of metalloproteinase I (TIMP-1), which acts to inhibit the activity of matrix metalloproteinases. The purpose of the present study was to determine how leptin, a gp130 cytokine, orchestrates the regulation of TIMP-1 gene activation and expression. Transient transfection of primary HSCs revealed that leptin significantly increased luciferase activity of a 229-bp TIMP-1 promoter construct (TIMP-1-229). An EMSA revealed that leptin enhanced specificity protein 1 (Sp1) binding. Site-directed mutagenesis for Sp1 reduced the enhancing effect of leptin on TIMP-1 transcriptional activation, and this effect was dose dependent on the number of Sp1 sites mutated. Chromatin immunoprecipitation revealed that leptin enhanced binding of Sp1; however, inhibition of signal transducer and activator of transcription (STAT) 3 phosphorylation by AG490 also blocked Sp1 phosphorylation and significantly reduced leptin-associated TIMP-1-229 promoter activity, indicating that one mechanism for leptin-increased transcriptional activity is via phosphorylation of Sp1 and subsequent promoter binding. Finally, we demonstrate that leptin also results in intranuclear pSTAT3 binding to Sp1. We propose a novel mechanism whereby leptin-mediated TIMP-1 transcription employs a Sp1/pSTAT3-dependent mechanism, one of which is a noncanonical association between Sp1 and pSTAT3. These data provide a new molecular mechanism whereby the adipocytokine leptin plays a role in complications of the metabolic syndrome.
Lin
Songbai
S
Department of Medicine, Division of Digestive Diseases, Emory University School of Medicine, Room 248, Whitehead Biomedical Research Building, 615 Michael Street, Atlanta, Georgia 30322, USA.
Saxena
Neeraj K
NK
Ding
Xiaokun
X
Stein
Lance L
LL
Anania
Frank A
FA
D000818:Animals; D001483:Base Sequence; D047369:Chromatin Immunoprecipitation; D015870:Gene Expression / Q000187:drug effects*; D005786:Gene Expression Regulation; D020738:Leptin / Q000494:pharmacology*; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D050796:STAT3 Transcription Factor / Q000378:metabolism*; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D051705:Sp3 Transcription Factor / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics*; D015533:Transcriptional Activation
D020738:Leptin; D009841:Oligonucleotides; D012333:RNA, Messenger; D050796:STAT3 Transcription Factor; D016329:Sp1 Transcription Factor; D019715:Tissue Inhibitor of Metalloproteinase-1; D051705:Sp3 Transcription Factor