Two domains of ISGF3 gamma that mediate protein-DNA and protein-protein interactions during transcription factor assembly contribute to DNA-binding specificity.
Alpha interferon (IFN-alpha) induces the transcription of a large set of genes through activation of multimeric transcription factor ISGF3. This factor can be dissociated into two protein components, termed ISGF3 gamma and ISGF3 alpha. ISGF3 gamma is a 48-kDa protein related at the amino terminus to members of the IFN-regulatory factor (IRF) and Myb families of DNA-binding proteins; ISGF3 alpha consists of three polypeptides of 84, 91, and 113 kDa that self-assemble to form an activated component in response to IFN-alpha. DNA-binding studies indicated that ISGF3 gamma binds DNA alone, recognizing the IFN-stimulated response element, while the ISGF3 alpha polypeptides alone display no specific interactions with DNA. A complex between ISGF3 gamma and activated ISGF3 alpha binds the IFN-stimulated response element with much greater affinity than does the 48-kDa ISGF3 gamma protein alone. The DNA-binding domain of ISGF3 gamma and regions responsible for protein-protein interaction with ISGF3 alpha were identified by using deleted forms of ISGF3 gamma expressed in vitro. The amino-terminal region of ISGF3 gamma homologous to the IRF and Myb proteins was sufficient for interaction with DNA and displayed the binding specificity of the intact protein; phosphorylation of this region was necessary for activity. A second region of 160 amino acids separated from the DNA-binding domain by over 100 amino acids contained a domain capable of associating with ISGF3 alpha and was sufficient to confer specific ISGF3 alpha interaction to a heterologous protein. Interaction of the ISGF3 alpha component with the protein interaction domain of ISGF3 gamma altered the DNA-binding specificity of the resulting complex, suggesting that one or more of the ISGF3 alpha polypeptides make base-specific contacts with DNA. This interaction defines a mechanism through which IRF-like proteins complexed with regulatory components can display novel DNA-binding specificities.
Veals
S A
SA
Department of Pathology, New York University School of Medicine, New York 10016.
Santa Maria
T
T
Levy
D E
DE
D001483:Base Sequence; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D050832:Interferon-Stimulated Gene Factor 3; D050833:Interferon-Stimulated Gene Factor 3, alpha Subunit; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism
D004268:DNA-Binding Proteins; C494227:IRF9 protein, human; D050832:Interferon-Stimulated Gene Factor 3; D050833:Interferon-Stimulated Gene Factor 3, alpha Subunit; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D014157:Transcription Factors