Insulin-activated Elk-1 recruits the TIP60/NuA4 complex to increase prolactin gene transcription.
Insulin increases prolactin gene expression in GH4 cells through phosphorylation of Elk-1 (Jacob and Stanley, 2001). We preformed a reverse two-hybrid screen using Elk-1-B42 as bait to identify proteins from GH4 cells that might serve as co-activators or co-repressors for insulin-increased prolactin gene expression. A number of the components of the TIP60/NuA4 complex interacted with Elk-1 suggesting that Elk-1 might activate transcription by recruiting the TIP60 chromatin-remodeling complex to the prolactin promoter. Inhibition of insulin-increased prolactin-luciferase expression by wild type and mutant adenovirus E1A protein provided physiological context for these yeast studies. Inhibition of histone deacetylases dramatically increased both basal and insulin-increased prolactin gene transcription. Co-immune precipitation experiments demonstrated Elk-1 and TIP60 associate in vitro. Transient or stable expression of TIP60 activated insulin-increased prolactin gene expression while a mutated TIP60 blocked insulin-increased prolactin gene expression. Analysis of the prolactin mRNA by quantitative RT-PCR showed that insulin-increased prolactin mRNA accumulation and that this was inhibited in GH4 cells that stably expressed mutant TIP60. Finally, ChIP experiments demonstrate the insulin-dependent occupancy of the prolactin promoter by Elk-1 and TIP60. Our studies suggest that insulin activates prolactin gene transcription by activating Elk-1 that recruits the NuA4 complex to the promoter.
Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY 10016, United States.
Muktar A
MA
Mahajan
Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY 10016, United States; NYU Cancer Institute, New York University School of Medicine, New York, NY 10016, United States. Electronic address: Frederick.Stanley@nyumc.org.
Frederick M
FM
Stanley
D000107:Acetylation / Q000187:drug effects; D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D002843:Chromatin / Q000187:drug effects / Q000378:metabolism; D042002:Chromatin Assembly and Disassembly / Q000187:drug effects; D047369:Chromatin Immunoprecipitation; D005786:Gene Expression Regulation / Q000187:drug effects*; D006657:Histones / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D008156:Luciferases / Q000378:metabolism; D050505:Mutant Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D011388:Prolactin / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D015534:Trans-Activators / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects*; D020798:Two-Hybrid System Techniques; D050807:ets-Domain Protein Elk-1 / Q000378:metabolism*; D050880:p300-CBP Transcription Factors / Q000378:metabolism
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D002843:Chromatin; C494114:Elk1 protein, rat; D006657:Histones; D007328:Insulin; D050505:Mutant Proteins; D009687:Nuclear Proteins; D015534:Trans-Activators; D050807:ets-Domain Protein Elk-1; C113829:transformation-transcription domain-associated protein; D011388:Prolactin; D008156:Luciferases; D050880:p300-CBP Transcription Factors
3,5,3′-triiodo-L-thyronine; Chromatin-remodeling; EGF; Elk-1; GH; HAT; HIR; Insulin; Luc; Prolactin; T3; TIP60; Transcription; epidermal growth factor; growth hormone; histone acetyltransferase; human insulin receptor; luciferase; β-gal; β-galactosidase