title: The 48 bp centromeric repeat is a functionally conserved motif in great apes and man showing protein-binding properties.
abstract: The centromere-kinetochore complex is a chromosomal assembly site including repeat motifs and protein binding properties thus mediating chromosome motility and mitotic regulation. Next to the alpha-satellite DNA family as well as human satellite III DNA, contribution of other repetitive sequences has to be strongly considered in centromere function. Here, we report the identification of centromeric 48 bp motifs, isolated from chimpanzee and orang-utan using an orthologous human DNA probe. Applying Southern hybridization we show that these sequences are restricted to hominoid species. Diminishing hybrid formation in gibbons suggested that the 48 bp repeat originated approximately 25-20 million years ago. Consistently, both chimpanzee as well as human repeat probes failed to generate any hybridization signal with the monkey species Cercopithecus aethiops and Aotes trivirgatus. Sequence deviations from the consensus of human repeat monomers of 6% and 10.4% in chimpanzee and orang-utan, respectively, were found within a 16 bp region of the 48 bp repeat units. Gel mobility shift assays using chimpanzee repeat dimers as probes revealed peptide-binding properties with human and chimpanzee nuclear extracts. Species-specific DNA-protein complexes remained unaffected by competition studies and indicated the presence of at least one novel interacting protein consisting of two subunits with 90 and 95 kDa. Our data suggest that the 48 bp repeat, next to alpha-satellite DNA, provides essential sequence information for specific DNA-protein interaction and they imply phylogenetic conservation of these binding properties in primates. The complex is likely involved in the proper formation and/or function of mammalian centromeres.
authors.affiliation: Institute of Anthropology and Human Genetics, Eberhard-Karls University, Tübingen, Germany. carsten.pusch@uni-tuebingen.de
authors.forename: Carsten M
authors.initials: CM
authors.lastname: Pusch
authors.forename: Iris
authors.initials: I
authors.lastname: Wundrack
authors.forename: Roman
authors.initials: R
authors.lastname: Müllenbach
authors.forename: Werner
authors.initials: W
authors.lastname: Schempp
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Blin
mesh_terms: D000818:Animals; D001483:Base Sequence; D026501:Blotting, Southwestern; D002503:Centromere / Q000235:genetics* / Q000378:metabolism; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000032:analysis / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D015186:Hominidae / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D011485:Protein Binding; D012091:Repetitive Sequences, Nucleic Acid; D016415:Sequence Alignment
chemical_list: D004268:DNA-Binding Proteins