title: Both tails and the centromere targeting domain of CENP-A are required for centromere establishment.
abstract: The centromere-defined by the presence of nucleosomes containing the histone H3 variant, CENP-A-is the chromosomal locus required for the accurate segregation of chromosomes during cell division. Although the sequence determinants of human CENP-A required to maintain a centromere were reported, those that are required for early steps in establishing a new centromere are unknown. In this paper, we used gain-of-function histone H3 chimeras containing various regions unique to CENP-A to investigate early events in centromere establishment. We targeted histone H3 chimeras to chromosomally integrated Lac operator sequences by fusing each of the chimeras to the Lac repressor. Using this approach, we found surprising contributions from a small portion of the N-terminal tail and the CENP-A targeting domain in the initial recruitment of two essential constitutive centromere proteins, CENP-C and CENP-T. Our results indicate that the regions of CENP-A required for early events in centromere establishment differ from those that are required for maintaining centromere identity.
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Glennis A
authors.initials: GA
authors.lastname: Logsdon
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3QR, UK Faculty of Biology, Albert Ludwigs Universität Freiburg, 79104 Freiburg, Germany.
authors.forename: Evelyne J
authors.initials: EJ
authors.lastname: Barrey
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Emily A
authors.initials: EA
authors.lastname: Bassett
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Jamie E
authors.initials: JE
authors.lastname: DeNizio
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Lucie Y
authors.initials: LY
authors.lastname: Guo
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Tanya
authors.initials: T
authors.lastname: Panchenko
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104.
authors.forename: Jennine M
authors.initials: JM
authors.lastname: Dawicki-McKenna
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3QR, UK.
authors.forename: Patrick
authors.initials: P
authors.lastname: Heun
authors.affiliation: Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 Department of Biochemistry and Biophysics, Graduate Program in Biochemistry and Molecular Biophysics, and Graduate Program in Cell and Molecular Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 blackbe@mail.med.upenn.edu.
authors.forename: Ben E
authors.initials: BE
authors.lastname: Black
mesh_terms: D001324:Autoantigens / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002503:Centromere / Q000235:genetics / Q000378:metabolism*; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism*; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D017434:Protein Structure, Tertiary
chemical_list: D001324:Autoantigens; C000619700:CENPA protein, human; C534001:CENPT protein, human; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone; D006657:Histones; C060905:centromere protein C