The Pit-1beta domain dictates active repression and alteration of histone acetylation of the proximal prolactin promoter.
A critical problem in current molecular biology is to gain a detailed understanding of the molecular mechanisms by which related transcription factor isoforms with identical DNA sequence specificity mediate distinct transcription responses. Pit-1 and Pit-1beta constitute such a pair of transcription factor isoforms. Pit-1 enhances the Ras signaling pathway to the prolactin promoter, and Pit-1beta represses basal prolactin promoter activity as well as Ras signaling to the prolactin promoter in pituitary cells. We have previously demonstrated that the beta-domain amino acid sequence dictates the transcriptional properties of Pit-1beta. Here, we show that five hydrophobic beta-domain residues are required for Pit-1 isoform-specific repression of Ras signaling, and we demonstrate that sodium butyrate and trichostatin A, pharmacological inhibitors of histone deacetylation, as well as viral Ski protein, a dominant-negative inhibitor of recruitment of N-CoR/mSin3 histone deacetylase complexes, specifically reverse beta isoform-specific repression of Ras signaling. Moreover, we directly demonstrate, with a chromatin immunoprecipitation assay, that the Pit-1beta isoform alters the histone acetylation state of the proximal prolactin promoter. This differential analysis of Pit-1/Pit-1beta isoform function provides significant insights into the structural determinants that govern how different transcription factors with identical DNA sequence specificity can display opposite effects on target gene activity.
Diamond
S E
SE
Department of Medicine and Department of Biochemistry and Molecular Genetics, Program in Molecular Biology and Colorado Cancer Center, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.
Gutierrez-Hartmann
A
A
D000107:Acetylation; D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002087:Butyrates / Q000494:pharmacology; D002843:Chromatin / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D004791:Enzyme Inhibitors; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D008156:Luciferases / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011388:Prolactin / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051381:Rats; D050817:Transcription Factor Pit-1; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D018631:ras Proteins / Q000378:metabolism
D002087:Butyrates; D002843:Chromatin; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D006657:Histones; D006877:Hydroxamic Acids; C494187:POU1F1 protein, human; C494186:Pou1f1 protein, rat; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; C060614:SKI protein, human; C012589:trichostatin A; D011388:Prolactin; D008156:Luciferases; D018631:ras Proteins; D000409:Alanine