A novel chromatin protein, distantly related to histone H2A, is largely excluded from the inactive X chromosome.
Chromatin on the mammalian inactive X chromosome differs in a number of ways from that on the active X. One protein, macroH2A, whose amino terminus is closely related to histone H2A, is enriched on the heterochromatic inactive X chromosome in female cells. Here, we report the identification and localization of a novel and more distant histone variant, designated H2A-Bbd, that is only 48% identical to histone H2A. In both interphase and metaphase female cells, using either a myc epitope-tagged or green fluorescent protein-tagged H2A-Bbd construct, the inactive X chromosome is markedly deficient in H2A-Bbd staining, while the active X and the autosomes stain throughout. In double-labeling experiments, antibodies to acetylated histone H4 show a pattern of staining indistinguishable from H2A-Bbd in interphase nuclei and on metaphase chromosomes. Chromatin fractionation demonstrates association of H2A-Bbd with the histone proteins. Separation of micrococcal nuclease-digested chromatin by sucrose gradient ultracentrifugation shows cofractionation of H2A-Bbd with nucleosomes, supporting the idea that H2A-Bbd is incorporated into nucleosomes as a substitute for the core histone H2A. This finding, in combination with the overlap with acetylated forms of H4, raises the possibility that H2A-Bbd is enriched in nucleosomes associated with transcriptionally active regions of the genome. The distribution of H2A-Bbd thus distinguishes chromatin on the active and inactive X chromosomes.
Department of Genetics, Case Western Reserve University School of Medicine and Center for Human Genetics and Research Institute, University Hospitals of Cleveland, Cleveland, Ohio 44106-4955, USA.
B P
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Chadwick
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Willard
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000235:genetics*; D002874:Chromosome Mapping; D020224:Expressed Sequence Tags; D005260:Female; D014644:Genetic Variation; D006570:Heterochromatin / Q000235:genetics; D006657:Histones / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007680:Kidney Neoplasms; D008322:Mammals; D008969:Molecular Sequence Data; D005810:Multigene Family; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D014960:X Chromosome
D002843:Chromatin; D006570:Heterochromatin; D006657:Histones; D011993:Recombinant Fusion Proteins