Modular structure of chromosomal proteins HMG-14 and HMG-17: definition of a transcriptional enhancement domain distinct from the nucleosomal binding domain.
Chromosomal proteins HMG-14 and HMG-17 are the only known nuclear proteins which specifically bind to the nucleosome core particle and are implicated in the generation and/or maintenance of structural features specific to active chromatin. The two proteins facilitate polymerase II and III transcription from in vitro- and in vivo-assembled circular chromatin templates. Here we used deletion mutants and specific peptides to identify the transcriptional enhancement domain and delineate the nucleosomal binding domain of the HMG-14 and -17 proteins. Deletion of the 22 C-terminal amino acids of HMG-17 or 26 C-terminal amino acids of HMG-14 reduces significantly the ability of the proteins to enhance transcription from chromatin templates. In contrast, N-terminal truncation mutants had the same transcriptional enhancement activity as the full-length proteins. We conclude that the negatively charged C-terminal region of the proteins is required for transcriptional enhancement. Chromatin transcription enhancement assays, which involve binding competition between the full-length proteins and peptides derived from their nucleosomal binding regions, indicate that the minimal nucleosomal binding domain of human HMG-17 is 24 amino acids long and spans residues 17 to 40. The results suggest that HMG-14 and -17 proteins have a modular structure and contain distinct functional domains.
Laboratory of Molecular Carcinogenesis, National Cancer Institute, Bethesda, Maryland 20892, USA.
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Trieschmann
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YV
Postnikov
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Rickers
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Bustin
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002843:Chromatin / Q000378:metabolism* / Q000648:ultrastructure*; D005260:Female; D006609:High Mobility Group Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009707:Nucleosomes / Q000378:metabolism*; D009865:Oocytes / Q000502:physiology; D010446:Peptide Fragments / Q000494:pharmacology; D012341:RNA, Ribosomal, 5S / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014981:Xenopus; D014982:Xenopus laevis
D002843:Chromatin; D006609:High Mobility Group Proteins; D009707:Nucleosomes; D010446:Peptide Fragments; D012341:RNA, Ribosomal, 5S; D011994:Recombinant Proteins