In vivo protein binding sites and nuclease hypersensitivity in the promoter region of a cell cycle regulated human H3 histone gene.
The chromatin structure and protein-DNA interactions of a cell cycle regulated human H3 histone gene have been examined at different levels of resolution. Using traditional Southern blot analysis we have investigated the accessibility of the H3 coding region and its flanking sequences to DNase I, S1 nuclease and restriction endonuclease digestion. Using the native genomic blotting method recently developed in our laboratory, two sites of protein-DNA interaction in the proximal 240 bp of the promoter region of this H3 gene were established. Further in vivo analysis of protein-DNA binding sites in intact cells by genomic sequencing revealed, with single nucleotide resolution, the guanine contacts and footprints of the proteins bound to the promoter. The relative locations of protein-DNA interactions in this H3 gene are similar to those identified in vivo and in vitro in a cell cycle dependent human H4 histone gene. The proteins complexed with the H3 histone gene promoter can be dissociated between 0.16 and 0.28 M NaCl. The protein-DNA contacts persist throughout the cell cycle and thus may have a functional relationship with the basal level of transcription of this H3 gene that occurs during and outside of S phase.
Pauli
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U
University of Florida, College of Medicine, Gainesville 32610.
Chrysogelos
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S
Nick
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H
Stein
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Stein
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J
D001483:Base Sequence; D001665:Binding Sites; D002453:Cell Cycle; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004720:Endonucleases / Q000378:metabolism; D006367:HeLa Cells; D006657:Histones / Q000235:genetics*; D006801:Humans; D007399:Interphase; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011506:Proteins / Q000378:metabolism*; D015719:Single-Strand Specific DNA and RNA Endonucleases; D012965:Sodium Chloride / Q000494:pharmacology; D014158:Transcription, Genetic
D006657:Histones; D011506:Proteins; D012965:Sodium Chloride; D004720:Endonucleases; D004262:DNA Restriction Enzymes; C052155:endodeoxyribonuclease MnlI; D003850:Deoxyribonuclease I; D015252:Deoxyribonucleases, Type II Site-Specific; D015719:Single-Strand Specific DNA and RNA Endonucleases; D010727:Phosphoric Diester Hydrolases