title: Histone modifications rather than the novel regional centromeres of Zymoseptoria tritici distinguish core and accessory chromosomes.
abstract: BACKGROUND
  Supernumerary chromosomes have been found in many organisms. In fungi, these "accessory" or "dispensable" chromosomes are present at different frequencies in populations and are usually characterized by higher repetitive DNA content and lower gene density when compared to the core chromosomes. In the reference strain of the wheat pathogen, Zymoseptoria tritici, eight discrete accessory chromosomes have been found. So far, no functional role has been assigned to these chromosomes; however, they have existed as separate entities in the karyotypes of Zymoseptoria species over evolutionary time. In this study, we addressed what-if anything-distinguishes the chromatin of accessory chromosomes from core chromosomes. We used chromatin immunoprecipitation combined with high-throughput sequencing ("ChIP-seq") of DNA associated with the centromere-specific histone H3, CENP-A (CenH3), to identify centromeric DNA, and ChIP-seq with antibodies against dimethylated H3K4, trimethylated H3K9 and trimethylated H3K27 to determine the relative distribution and proportion of euchromatin, obligate and facultative heterochromatin, respectively.
  
  
  RESULTS
  Centromeres of the eight accessory chromosomes have the same sequence composition and structure as centromeres of the 13 core chromosomes and they are of similar length. Unlike those of most other fungi, Z. tritici centromeres are not composed entirely of repetitive DNA; some centromeres contain only unique DNA sequences, and bona fide expressed genes are located in regions enriched with CenH3. By fluorescence microscopy, we showed that centromeres of Z. tritici do not cluster into a single chromocenter during interphase. We found dramatically higher enrichment of H3K9me3 and H3K27me3 on the accessory chromosomes, consistent with the twofold higher proportion of repetitive DNA and poorly transcribed genes. In contrast, no single histone modification tested here correlated with the distribution of centromeric nucleosomes.
  
  
  CONCLUSIONS
  All centromeres are similar in length and composed of a mixture of unique and repeat DNA, and most contain actively transcribed genes. Centromeres, subtelomeric regions or telomere repeat length cannot account for the differences in transfer fidelity between core and accessory chromosomes, but accessory chromosomes are greatly enriched in nucleosomes with H3K27 trimethylation. Genes on accessory chromosomes appear to be silenced by trimethylation of H3K9 and H3K27.
authors.affiliation: Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch Strasse 10, 35043 Marburg, Germany ; Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR 97331-7303 USA ; Christian-Albrechts University of Kiel, Environmental Genomics, Am Botanischen Garten 9-11, 24118 Kiel, Germany ; Max Planck Institute for Evolutionary Biology, August-Thienemann-Str. 2, 24306 Plön, Germany.
authors.forename: Klaas
authors.initials: K
authors.lastname: Schotanus
authors.affiliation: Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch Strasse 10, 35043 Marburg, Germany ; INRA, UMR 1290 INRA-AgroParisTech BIOGER, Avenue Lucien Brétignières, Thiverval-Grignon, 78850 France ; Christian-Albrechts University of Kiel, Environmental Genomics, Am Botanischen Garten 9-11, 24118 Kiel, Germany ; Max Planck Institute for Evolutionary Biology, August-Thienemann-Str. 2, 24306 Plön, Germany.
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Soyer
authors.affiliation: Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR 97331-7303 USA.
authors.forename: Lanelle R
authors.initials: LR
authors.lastname: Connolly
authors.affiliation: Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch Strasse 10, 35043 Marburg, Germany ; Christian-Albrechts University of Kiel, Environmental Genomics, Am Botanischen Garten 9-11, 24118 Kiel, Germany ; Max Planck Institute for Evolutionary Biology, August-Thienemann-Str. 2, 24306 Plön, Germany.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Grandaubert
authors.affiliation: Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch Strasse 10, 35043 Marburg, Germany.
authors.forename: Petra
authors.initials: P
authors.lastname: Happel
authors.affiliation: Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR 97331-7303 USA.
authors.forename: Kristina M
authors.initials: KM
authors.lastname: Smith
authors.affiliation: Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR 97331-7303 USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Freitag
authors.affiliation: Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch Strasse 10, 35043 Marburg, Germany ; Christian-Albrechts University of Kiel, Environmental Genomics, Am Botanischen Garten 9-11, 24118 Kiel, Germany ; Max Planck Institute for Evolutionary Biology, August-Thienemann-Str. 2, 24306 Plön, Germany.
authors.forename: Eva H
authors.initials: EH
authors.lastname: Stukenbrock
keywords: Accessory chromosomes; Centromere; ChIP-seq; Histone methylation; Zymoseptoria tritici (Mycosphaerella graminicola)