The centromere is the chromosome region where microtubules attach during cell division. In contrast to monocentric chromosomes with one centromere, holocentric species usually distribute hundreds of centromere units along the entire chromatid. We assembled the chromosome-scale reference genome and analyzed the holocentromere and (epi)genome organization of the lilioid Chionographis japonica. Remarkably, each of its holocentric chromatids consists of only 7 to 11 evenly spaced megabase-sized centromere-specific histone H3-positive units. These units contain satellite arrays of 23 and 28 bp-long monomers capable of forming palindromic structures. Like monocentric species, C. japonica forms clustered centromeres in chromocenters at interphase. In addition, the large-scale eu- and heterochromatin arrangement differs between C. japonica and other known holocentric species. Finally, using polymer simulations, we model the formation of prometaphase line-like holocentromeres from interphase centromere clusters. Our findings broaden the knowledge about centromere diversity, showing that holocentricity is not restricted to species with numerous and small centromere units.

Arid Land Research Center Tottori University 1390 Hamasaka Tottori 680 0001 Japan

Biology Centre Czech Academy of Sciences Institute of Plant Molecular Biology Branišovská 31 České Budějovice CZ 37005 Czech Republic

Department of Chromosome Biology Max Planck Institute for Plant Breeding Research Cologne 50829 Germany

Julius Kühn Institute Institute for Breeding Research on Horticultural Crops Erwin Baur Str 27 06484 Quedlinburg Germany

Leibniz Institute of Plant Genetics and Crop Plant Research Gatersleben Corrensstrasse 3 06466 Seeland Germany

Max Planck Genome Centre Cologne Max Planck Institute for Plant Breeding Research Cologne 50829 Germany

Zobrazit více v PubMed

Talbert PB, Henikoff S. What makes a centromere? Exp. Cell Res. 2020;389:111895. doi: 10.1016/j.yexcr.2020.111895. PubMed DOI

Muller H, Gil J, Jr, Drinnenberg IA. The impact of centromeres on spatial genome architecture. Trends Genet. 2019;35:565–578. doi: 10.1016/j.tig.2019.05.003. PubMed DOI

Buchwitz BJ, Ahmad K, Moore LL, Roth MB, Henikoff S. Cell division - A histone-H3-like protein in C. elegans. Nature. 1999;401:547–548. doi: 10.1038/44062. PubMed DOI

Nagaki K, Kashihara K, Murata M. Visualization of diffuse centromeres with centromere-specific histone H3 in the holocentric plant Luzula nivea. Plant Cell. 2005;17:1886–1893. doi: 10.1105/tpc.105.032961. PubMed DOI PMC

Marques A, et al. Holocentromeres in Rhynchospora are associated with genome-wide centromere-specific repeat arrays interspersed among euchromatin. Proc. Natl Acad. Sci. USA. 2015;112:13633–13638. doi: 10.1073/pnas.1512255112. PubMed DOI PMC

Schrader F. The role of the kinetochore in the chromosomal evolution of the Heteroptera and Homoptera. Evolution. 1947;1:134–142. doi: 10.2307/2405489. DOI

Camara AS, Schubert V, Mascher M, Houben A. A simple model explains the cell cycle-dependent assembly of centromeric nucleosomes in holocentric species. Nucleic Acids Res. 2021;49:9053–9065. doi: 10.1093/nar/gkab648. PubMed DOI PMC

Melters DP, Paliulis LV, Korf IF, Chan SW. Holocentric chromosomes: convergent evolution, meiotic adaptations, and genomic analysis. Chromosome Res. 2012;20:579–593. doi: 10.1007/s10577-012-9292-1. PubMed DOI

Senaratne AP, Cortes-Silva N, Drinnenberg IA. Evolution of holocentric chromosomes: Drivers, diversity, and deterrents. Semin Cell Dev. Biol. 2022;127:90–99. doi: 10.1016/j.semcdb.2022.01.003. PubMed DOI

Grzan T, Despot-Slade E, Mestrovic N, Plohl M, Mravinac B. CenH3 distribution reveals extended centromeres in the model beetle Tribolium castaneum. PLoS Genet. 2020;16:e1009115. doi: 10.1371/journal.pgen.1009115. PubMed DOI PMC

Neumann P, et al. Centromeres off the hook: massive changes in centromere size and structure following duplication of CenH3 gene in Fabeae species. Mol. Biol. Evol. 2015;32:1862–1879. doi: 10.1093/molbev/msv070. PubMed DOI PMC

Schubert V, et al. Super-resolution microscopy reveals diversity of plant centromere architecture. Int. J. Mol. Sci. 2020;21:3488. doi: 10.3390/ijms21103488. PubMed DOI PMC

Drinnenberg IA, deYoung D, Henikoff S, Malik HS. Recurrent loss of CenH3 is associated with independent transitions to holocentricity in insects. Elife. 2014;3:e03676. doi: 10.7554/eLife.03676. PubMed DOI PMC

Slade ED, et al. The centromere histone is conserved and associated with tandem repeats sharing a conserved 19 bp box in the holocentromere of Meloidogyne nematodes. Mol. Biol. Evol. 2021;38:1943–1965. doi: 10.1093/molbev/msaa336. PubMed DOI PMC

Oliveira L, et al. Mitotic spindle attachment to the holocentric chromosomes of Cuscuta europaea does not correlate with the distribution of CENH3 chromatin. Front. Plant Sci. 2019;10:1799. doi: 10.3389/fpls.2019.01799. PubMed DOI PMC

Senaratne AP, et al. Formation of the CenH3-deficient holocentromere in Lepidoptera avoids active chromatin. Curr. Biol. 2021;31:173. doi: 10.1016/j.cub.2020.09.078. PubMed DOI

Hofstatter PG, et al. Repeat-based holocentromeres influence genome architecture and karyotype evolution. Cell. 2022;185:3153–3168.e3118. doi: 10.1016/j.cell.2022.06.045. PubMed DOI

Tanaka N, Tanaka N. Chromosome studies in Chionographis (Liliaceae). I. On the holokintic nature of chromosomes in Chionographis japonica Maxim. Cytologica. 1977;42:753–763. doi: 10.1508/cytologia.42.753. DOI

Tanaka N. High stability in chromosomal traits of Chamaelirium japonicum and C. koidzuminum (Melanthiaceae) with holocentric chromosomes. Cytologia. 2020;85:33–40. doi: 10.1508/cytologia.85.33. DOI

Tanaka N, Tanaka N. Chromosome studies in Chionographis (Liliaceae). 2. Morphological characteristics of the somatic chromosomes of 4 Japanese members. Cytologia. 1979;44:935–949. doi: 10.1508/cytologia.44.935. DOI

Wanner G, Schroeder-Reiter E, Ma W, Houben A, Schubert V. The ultrastructure of mono- and holocentric plant centromeres: an immunological investigation by structured illumination microscopy and scanning electron microscopy. Chromosoma. 2015;124:503–517. doi: 10.1007/s00412-015-0521-1. PubMed DOI

Fransz P, de Jong JH, Lysak M, Castiglione MR, Schubert I. Interphase chromosomes in Arabidopsis are organized as well defined chromocenters from which euchromatin loops emanate. Proc. Natl Acad. Sci. USA. 2002;99:14584–14589. doi: 10.1073/pnas.212325299. PubMed DOI PMC

Du Y, Dawe RK. Maize NDC80 is a constitutive feature of the central kinetochore. Chromosome Res. 2007;15:767–775. doi: 10.1007/s10577-007-1160-z. PubMed DOI

Sato H, Shibata F, Murata M. Characterization of a Mis12 homologue in Arabidopsis thaliana. Chromosome Res. 2005;13:827–834. doi: 10.1007/s10577-005-1016-3. PubMed DOI

Hindriksen S, Lens SMA, Hadders MA. The Ins and Outs of Aurora B Inner Centromere Localization. Front. Cell Dev. Biol. 2017;5:112. doi: 10.3389/fcell.2017.00112. PubMed DOI PMC

Gernand D, Demidov D, Houben A. The temporal and spatial pattern of histone H3 phosphorylation at serine 28 and serine 10 is similar in plants but differs between mono- and polycentric chromosomes. Cytogenet. Genome Res. 2003;101:172–176. doi: 10.1159/000074175. PubMed DOI

Demidov D, et al. Anti-phosphorylated histone H2AThr120: a universal microscopic marker for centromeric chromatin of mono- and holocentric plant species. Cytogenet. Genome Res. 2014;143:150–156. doi: 10.1159/000360018. PubMed DOI

Novak P, Neumann P, Macas J. Global analysis of repetitive DNA from unassembled sequence reads using RepeatExplorer2. Nat. Protoc. 2020;15:3745–3776. doi: 10.1038/s41596-020-0400-y. PubMed DOI

Heckmann S, et al. Holocentric chromosomes of Luzula elegans are characterized by a longitudinal centromere groove, chromosome bending, and a terminal nucleolus organizer region. Cytogenet. Genome Res. 2011;134:220–228. doi: 10.1159/000327713. PubMed DOI

Heckmann S, et al. The holocentric species Luzula elegans shows interplay between centromere and large-scale genome organization. Plant J. 2013;73:555–565. doi: 10.1111/tpj.12054. PubMed DOI

Houben A, et al. Methylation of histone H3 in euchromatin of plant chromosomes depends on basic nuclear DNA content. Plant J. 2003;33:967–973. doi: 10.1046/j.1365-313X.2003.01681.x. PubMed DOI

Rea S, et al. Regulation of chromatin structure by site-specific histone H3 methyltransferases. Nature. 2000;406:593–599. doi: 10.1038/35020506. PubMed DOI

Naish M, et al. The genetic and epigenetic landscape of the Arabidopsis centromeres. Science. 2021;374:eabi7489. doi: 10.1126/science.abi7489. PubMed DOI PMC

Tek AL, Kashihara K, Murata M, Nagaki K. Functional centromeres in Astragalus sinicus include a compact centromere-specific histone H3 and a 20-bp tandem repeat. Chromosome Res. 2011;19:969–978. doi: 10.1007/s10577-011-9247-y. PubMed DOI

Altemose N, et al. Complete genomic and epigenetic maps of human centromeres. Science. 2022;376:eabl4178. doi: 10.1126/science.abl4178. PubMed DOI PMC

Higgins AW, Gustashaw KM, Willard HF. Engineered human dicentric chromosomes show centromere plasticity. Chromosome Res. 2005;13:745–762. doi: 10.1007/s10577-005-1009-2. PubMed DOI

Steiner FA, Henikoff S. Holocentromeres are dispersed point centromeres localized at transcription factor hotspots. Elife. 2014;3:e02025. doi: 10.7554/eLife.02025. PubMed DOI PMC

Jagannathan, M., Cummings, R. & Yamashita, Y. M. A conserved function for pericentromeric satellite DNA. Elife7, 10.7554/eLife.34122 (2018). PubMed PMC

Neumann P, et al. Stretching the rules: monocentric chromosomes with multiple centromere domains. PLoS Genet. 2012;8:e1002777. doi: 10.1371/journal.pgen.1002777. PubMed DOI PMC

Tanaka N. Chromosomal traits of Chamaelirium luteum (Melanthiaceae) with particular focus on the large heterochromatic centromeres. Taiwania. 2020;65:286–294.

Kim C, Kim SC, Kim JH. Historical biogeography of Melanthiaceae: a case of out-of-North America through the Bering land bridge. Front. Plant Sci. 2019;10:396. doi: 10.3389/fpls.2019.00396. PubMed DOI PMC

Menzel G, et al. Diversity of a complex centromeric satellite and molecular characterization of dispersed sequence families in sugar beet (Beta vulgaris) Ann. Bot. 2008;102:521–530. doi: 10.1093/aob/mcn131. PubMed DOI PMC

Sproul JS, et al. Dynamic evolution of euchromatic satellites on the X chromosome in Drosophila melanogaster and the simulans clade. Mol. Biol. Evol. 2020;37:2241–2256. doi: 10.1093/molbev/msaa078. PubMed DOI PMC

Cohen S, Segal D. Extrachromosomal circular DNA in eukaryotes: possible involvement in the plasticity of tandem repeats. Cytogenet. Genome Res. 2009;124:327–338. doi: 10.1159/000218136. PubMed DOI

Navratilova A, Koblizkova A, Macas J. Survey of extrachromosomal circular DNA derived from plant satellite repeats. BMC Plant Biol. 2008;8:90. doi: 10.1186/1471-2229-8-90. PubMed DOI PMC

Zeitlin, S. G. et al. Double-strand DNA breaks recruit the centromeric histone CENP-A. Proc. Natl. Acad. Sci. USA. 106, 15762–15767 (2009). PubMed PMC

Cuacos M, H Franklin FC, Heckmann S. Atypical centromeres in plants-what they can tell us. Front. Plant Sci. 2015;6:913. doi: 10.3389/fpls.2015.00913. PubMed DOI PMC

Murashige T, Skoog F. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol. Plant. 1962;15:473–497. doi: 10.1111/j.1399-3054.1962.tb08052.x. DOI

Dolezel J, Bartos J, Voglmayr H, Greilhuber J. Nuclear DNA content and genome size of trout and human. Cytom. A. 2003;51:127–128, author reply 129. PubMed

Dolezel J, Binarova P, Lucretti S. Analysis of nuclear DNA content in plant cells by flow cytometry. Biol. Plant. 1989;31:113–120. doi: 10.1007/BF02907241. DOI

Andrews, S. FastQC A. Quality Control tool for High Throughput Sequence Data. http://www.bioinformatics.babraham.ac.uk/projects/fastqc (accessed on 24 November 2010).

Neumann P, Novak P, Hostakova N, Macas J. Systematic survey of plant LTR-retrotransposons elucidates phylogenetic relationships of their polyprotein domains and provides a reference for element classification. Mob. DNA. 2019;10:1. doi: 10.1186/s13100-018-0144-1. PubMed DOI PMC

Haas BJ, et al. De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nat. Protoc. 2013;8:1494–1512. doi: 10.1038/nprot.2013.084. PubMed DOI PMC

Grabherr MG, et al. Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat. Biotechnol. 2011;29:644–652. doi: 10.1038/nbt.1883. PubMed DOI PMC

Padmarasu S, Himmelbach A, Mascher M, Stein N. In situ Hi-C for plants: an improved method to detect long-range chromatin interactions. Methods Mol. Biol. 2019;1933:441–472. doi: 10.1007/978-1-4939-9045-0_28. PubMed DOI

Cheng HY, Concepcion GT, Feng XW, Zhang HW, Li H. Haplotype-resolved de novo assembly using phased assembly graphs with hifiasm. Nat. Methods. 2021;18:170. doi: 10.1038/s41592-020-01056-5. PubMed DOI PMC

Seppey M, Manni M, Zbodnov EM. BUSCO: assessing genome assembly and annotation completeness. Methods Mol. Biol. 2019;1962:227–245. doi: 10.1007/978-1-4939-9173-0_14. PubMed DOI

Manni M, Berkeley MR, Seppey M, Simao FA, Zdobnov EM. BUSCO Update: novel and streamlined workflows along with broader and deeper phylogenetic coverage for scoring of eukaryotic, prokaryotic, and viral genomes. Mol. Biol. Evol. 2021;38:4647–4654. doi: 10.1093/molbev/msab199. PubMed DOI PMC

Li H, et al. 1000 Genome Project Data Processing Subgroup (2009) The Sequence alignment/map (SAM) format and SAMtools. Bioinformatics. 2009;25:2078–2079. doi: 10.1093/bioinformatics/btp352. PubMed DOI PMC

Ghurye J, et al. Integrating Hi-C links with assembly graphs for chromosome-scale assembly. PLoS Comput. Biol. 2019;15:e1007273. doi: 10.1371/journal.pcbi.1007273. PubMed DOI PMC

Novak P, et al. TAREAN: a computational tool for identification and characterization of satellite DNA from unassembled short reads. Nucleic Acids Res. 2017;45:e111. doi: 10.1093/nar/gkx257. PubMed DOI PMC

Richards EJ, Ausubel FM. Isolation of a higher eukaryotic telomere from Arabidopsis thaliana. Cell. 1988;53:127–136. doi: 10.1016/0092-8674(88)90494-1. PubMed DOI

Gerlach WL, Bedbrook JR. Cloning and characterization of ribosomal RNA genes from wheat and barley. Nucleic Acids Res. 1979;7:1869–1885. doi: 10.1093/nar/7.7.1869. PubMed DOI PMC

Kuo YT, Hsu HL, Yeh CH, Chang SB. Application of a modified drop method for high-resolution pachytene chromosome spreads in two Phalaenopsis species. Mol. Cytogenet. 2016;9:44. doi: 10.1186/s13039-016-0254-8. PubMed DOI PMC

Krueger F, Andrews SR. Bismark: a flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics. 2011;27:1571–1572. doi: 10.1093/bioinformatics/btr167. PubMed DOI PMC

Lopez-Delisle L, et al. pyGenomeTracks: reproducible plots for multivariate genomic datasets. Bioinformatics. 2021;37:422–423. doi: 10.1093/bioinformatics/btaa692. PubMed DOI PMC

Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat. Methods. 2012;9:357–359. doi: 10.1038/nmeth.1923. PubMed DOI PMC

Tarasov A, Vilella AJ, Cuppen E, Nijman IJ, Prins P. Sambamba: fast processing of NGS alignment formats. Bioinformatics. 2015;31:2032–2034. doi: 10.1093/bioinformatics/btv098. PubMed DOI PMC

Zhang Y, et al. Model-based analysis of ChIP-Seq (MACS) Genome Biol. 2008;9:R137. doi: 10.1186/gb-2008-9-9-r137. PubMed DOI PMC

Stovner EB, Saetrom P. epic2 efficiently finds diffuse domains in ChIP-seq data. Bioinformatics. 2019;35:4392–4393. doi: 10.1093/bioinformatics/btz232. PubMed DOI

Ramirez F, et al. deepTools2: a next generation web server for deep-sequencing data analysis. Nucleic Acids Res. 2016;44:W160–W165. doi: 10.1093/nar/gkw257. PubMed DOI PMC

Eastman P, et al. OpenMM 7: rapid development of high performance algorithms for molecular dynamics. PLoS Comput. Biol. 2017;13:e1005659. doi: 10.1371/journal.pcbi.1005659. PubMed DOI PMC

Schrodinger, L. The PyMOL molecular graphics system. Version1, 0. (2010).

Repeat-based holocentromeres of the woodrush Luzula sylvatica reveal insights into the evolutionary transition to holocentricity

KNL1 and NDC80 represent new universal markers for the detection of functional centromeres in plants