title: The CENP-A N-tail confers epigenetic stability to centromeres via the CENP-T branch of the CCAN in fission yeast.
abstract: In most eukaryotes, centromeres are defined epigenetically by presence of the histone H3 variant CENP-A [1-3]. CENP-A-containing chromatin recruits the constitutive centromere-associated network (CCAN) of proteins, which in turn directs assembly of the outer kinetochore to form microtubule attachments and ensure chromosome segregation fidelity [4-6]. Whereas the mechanisms that load CENP-A at centromeres are being elucidated, the functions of its divergent N-terminal tail remain enigmatic [7-12]. Here, we employ the well-studied fission yeast centromere [13-16] to investigate the function of the CENP-A (Cnp1) N-tail. We show that alteration of the N-tail does not affect Cnp1 loading at centromeres, outer kinetochore formation, or spindle checkpoint signaling but nevertheless elevates chromosome loss. N-tail mutants exhibited synthetic lethality with an altered centromeric DNA sequence, with rare survivors harboring chromosomal fusions in which the altered centromere was epigenetically inactivated. Elevated centromere inactivation was also observed for N-tail mutants with unaltered centromeric DNA sequences. N-tail mutants specifically reduced localization of the CCAN proteins Cnp20/CENP-T and Mis6/CENP-I, but not Cnp3/CENP-C. Overexpression of Cnp20/CENP-T suppressed defects in an N-tail mutant, suggesting a link between reduced CENP-T recruitment and the observed centromere inactivation phenotype. Thus, the Cnp1 N-tail promotes epigenetic stability of centromeres in fission yeast, at least in part via recruitment of the CENP-T branch of the CCAN.
authors.affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA; Laboratory of Biochemistry and Molecular Biology, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: H Diego
authors.initials: HD
authors.lastname: Folco
authors.affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Christopher S
authors.initials: CS
authors.lastname: Campbell
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Karen M
authors.initials: KM
authors.lastname: May
authors.affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Celso A
authors.initials: CA
authors.lastname: Espinoza
authors.affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA.
authors.forename: Karen
authors.initials: K
authors.lastname: Oegema
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
authors.forename: Kevin G
authors.initials: KG
authors.lastname: Hardwick
authors.affiliation: Laboratory of Biochemistry and Molecular Biology, National Cancer Institute, NIH, Bethesda, MD 20892, USA.
authors.forename: Shiv I S
authors.initials: SIS
authors.lastname: Grewal
authors.affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: abdesai@ucsd.edu.
authors.forename: Arshad
authors.initials: A
authors.lastname: Desai
mesh_terms: D002503:Centromere / Q000378:metabolism / Q000502:physiology*; D047369:Chromatin Immunoprecipitation; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics* / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D016521:Electrophoresis, Gel, Pulsed-Field; D044127:Epigenesis, Genetic / Q000502:physiology*; D005453:Fluorescence; D006657:Histones / Q000378:metabolism; D015151:Immunoblotting; D009154:Mutation / Q000235:genetics; D016133:Polymerase Chain Reaction; D012568:Schizosaccharomyces; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D002868:Chromosomal Proteins, Non-Histone; C410349:Cnp1 protein, S pombe; D017931:DNA Primers; D006657:Histones; D029702:Schizosaccharomyces pombe Proteins