Transcriptional inhibition by Stat5. Differential activities at growth-related versus differentiation-specific promoters.
Prolactin (PRL) induces transcriptional activation of not only growth-related genes such as interferon regulatory factor-1 (IRF-1) but also differentiation-specific genes such as beta-casein through a signaling cascade consisting of Janus kinases and Stat (signal transducer and activator of transcription) factors. To understand better the role of Stats in PRL signaling, we cloned rat Stat5b from a PRL-responsive T cell line Nb2. A Stat5b-specific peptide antibody was generated. In PRL receptor reconstituted COS cells cotransfected with Stat5b or Stat5a, both Stat5 proteins become tyrosine phosphorylated and bind to the IRF-1 GAS (interferon-gamma activation sequence) element in a PRL-inducible manner. Unexpectedly, both Stat5b and Stat5a inhibit PRL induction of the IRF-1 promoter, but they mediate PRL stimulation of the beta-casein promoter. Stat5-mediated inhibition was observed only at the native IRF-1 promoter and not at the isolated IRF-1 GAS element linked to a heterologous thymidine kinase promoter. Mutational analyses showed that the DNA binding activity of Stat5b is not required, but the carboxyl-terminal transactivation domain is essential for Stat5b to inhibit PRL induction of the IRF-1 promoter. These results suggest that Stat5b mediates inhibition via protein-protein interactions. In contrast, both DNA binding and transactivation domains of Stat5b are required to mediate PRL induction of the beta-casein promoter. Furthermore, a carboxyl-terminal truncated dominant negative Stat5b can reverse Stat5b inhibition at the IRF-1 promoter. These studies suggest that Stat proteins can act as not only positive but also negative regulators of gene transcription. Further, Stat5 can modulate gene expression without binding to DNA but via protein-protein interactions.
Luo
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Department of Microbiology and Immunology, Baylor College of Medicine, Houston, Texas 77030, USA.
Yu-Lee
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D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002454:Cell Differentiation; D002455:Cell Division; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D050856:Interferon Regulatory Factor-1; D016399:Lymphoma, T-Cell; D008894:Milk Proteins; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011388:Prolactin / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D051381:Rats; D011981:Receptors, Prolactin / Q000096:biosynthesis / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D050799:STAT5 Transcription Factor; D015534:Trans-Activators / Q000096:biosynthesis / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins
D004268:DNA-Binding Proteins; C494257:IRF1 protein, human; D050856:Interferon Regulatory Factor-1; C494259:Irf1 protein, rat; D008894:Milk Proteins; D010750:Phosphoproteins; D011981:Receptors, Prolactin; D011994:Recombinant Proteins; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; C494095:STAT5B protein, human; C494094:Stat5a protein, rat; C494097:Stat5b protein, rat; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D011388:Prolactin