The segregation of chromosomes depends on the centromere. Most species are monocentric, with the centromere restricted to a single region per chromosome. In some organisms, the monocentric organization changed to holocentric, in which the centromere activity is distributed over the entire chromosome length. However, the causes and consequences of this transition are poorly understood. Here, we show that the transition in the genus Cuscuta was associated with dramatic changes in the kinetochore, a protein complex that mediates the attachment of chromosomes to microtubules. We found that in holocentric Cuscuta species, the KNL2 genes were lost; the CENP-C, KNL1, and ZWINT1 genes were truncated; the centromeric localization of CENH3, CENP-C, KNL1, MIS12, and NDC80 proteins was disrupted; and the spindle assembly checkpoint (SAC) degenerated. Our results demonstrate that holocentric Cuscuta species lost the ability to form a standard kinetochore and do not employ SAC to control the attachment of microtubules to chromosomes.

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

Department of Biological Science College of Bioscience and Biotechnology Chungnam National University Daejeon 34134 Republic of Korea

Department of Breeding Research Leibniz Institute of Plant Genetics and Crop Plant Research Gatersleben 06466 Seeland Germany

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Melters D. P., Paliulis L. V., Korf I. F., Chan S. W. L., Holocentric chromosomes: Convergent evolution, meiotic adaptations, and genomic analysis. Chromosome Res. 20, 579–93 (2012). PubMed

McKinley K. L., Cheeseman I. M., The molecular basis for centromere identity and function. Nat. Rev. Mol. Cell Biol. 17, 16–29 (2016). PubMed PMC

Pesenti M. E., Weir J. R., Musacchio A., Progress in the structural and functional characterization of kinetochores. Curr. Opin. Struct. Biol. 37, 152–163 (2016). PubMed

Yamagishi Y., Sakuno T., Goto Y., Watanabe Y., Kinetochore composition and its function: Lessons from yeasts. FEMS Microbiol. Rev. 38, 185–200 (2014). PubMed

Lara-Gonzalez P., Westhorpe F. G., Taylor S. S., The spindle assembly checkpoint. Curr. Biol. 22, R966–R980 (2012). PubMed

Musacchio A., The molecular biology of spindle assembly checkpoint signaling dynamics. Curr. Biol. 25, R1002–R1018 (2015). PubMed

Komaki S., et al. , Functional analysis of the plant chromosomal passenger complex. Plant Physiol. 183, 1586–1599 (2020). PubMed PMC

Carmena M., Wheelock M., Funabiki H., Earnshaw W. C., The chromosomal passenger complex (CPC): From easy rider to the godfather of mitosis. Nat. Rev. Mol. Cell Biol. 13, 789–803 (2012). PubMed PMC

van der Waal M. S., Hengeveld R. C. C., van der Horst A., Lens S. M. A., Cell division control by the chromosomal passenger complex. Exp. Cell Res. 318, 1407–1420 (2012). PubMed

Buchwitz B. J., Ahmad K., Moore L. L., Roth M. B., Henikoff S., A histone-H3-like protein in C. elegans. Nature 401, 547–548 (1999). PubMed

Schubert V., et al. , Super-resolution microscopy reveals diversity of plant centromere architecture. Int. J. Mol. Sci. 21, 3488 (2020). PubMed PMC

Cortes-Silva N., et al. , CenH3-independent kinetochore assembly in Lepidoptera requires CCAN, including CENP-T. Curr. Biol. 30, 561–572.e10 (2020). PubMed

Drinnenberg I. A., DeYoung D., Henikoff S., Malik H. S., Recurrent loss of CenH3 is associated with independent transitions to holocentricity in insects. Elife 3, e03676 (2014). PubMed PMC

Senaratne A. P., et al. , Formation of the CenH3-deficient holocentromere in Lepidoptera avoids active chromatin. Curr. Biol. 31, 173–181.e7 (2021). PubMed

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. 10, 1799 (2020). PubMed PMC

Neumann P., et al. , Impact of parasitic lifestyle and different types of centromere organization on chromosome and genome evolution in the plant genus Cuscuta. New Phytol. 229, 2365–2377 (2021). PubMed

Hara M., Fukagawa T., Where is the right path heading from the centromere to spindle microtubules? Cell Cycle 18, 1199–1211 (2019). PubMed PMC

Petrovic A., et al. , Structure of the MIS12 complex and molecular basis of its interaction with CENP-C at human kinetochores. Cell 167, 1028–1040.e15 (2016). PubMed PMC

Ciferri C., et al. , Implications for kinetochore-microtubule attachment from the structure of an engineered Ndc80 complex. Cell 133, 427–439 (2008). PubMed PMC

Alushin G. M., et al. , The Ndc80 kinetochore complex forms oligomeric arrays along microtubules. Nature 467, 805–810 (2010). PubMed PMC

Welburn J. P. I., et al. , Aurora B phosphorylates spatially distinct targets to differentially regulate the kinetochore-microtubule interface. Mol. Cell 38, 383–392 (2010). PubMed PMC

Petrovic A., et al. , Modular assembly of RWD domains on the Mis12 complex underlies outer kinetochore organization. Mol. Cell 53, 591–605 (2014). PubMed

Valverde R., Ingram J., Harrison S. C., Conserved tetramer junction in the kinetochore Ndc80 complex. Cell Rep. 17, 1915–1922 (2016). PubMed PMC

Ali-Ahmad A., Bilokapić S., Schäfer I. B., Halić M., Sekulić N., CENP-C unwraps the human CENP-A nucleosome through the H2A C-terminal tail. EMBO Rep. 20, 1–13 (2019). PubMed PMC

Zuo S., et al. , Recurrent plant-specific duplications of KNL2 and its conserved function as a kinetochore assembly factor. Mol. Biol. Evol. 39 (2022). PubMed PMC

Hornung P., et al. , A cooperative mechanism drives budding yeast kinetochore assembly downstream of CENP-A. J. Cell Biol. 206, 509–524 (2014). PubMed PMC

Komaki S., Schnittger A., The spindle assembly checkpoint in Arabidopsis is rapidly shut off during severe stress. Dev. Cell 43, 172–185.e5 (2017). PubMed

Ghongane P., Kapanidou M., Asghar A., Elowe S., Bolanos-Garcia V. M., The dynamic protein Knl1 - a kinetochore rendezvous. J. Cell Sci. 127, 3415–3423 (2014). PubMed

Su H., et al. , Knl1 participates in spindle assembly checkpoint signaling in maize. Proc. Natl. Acad. Sci. U.S.A. 118, e2022357118 (2021). PubMed PMC

Caillaud M. C., et al. , Spindle assembly checkpoint protein dynamics reveal conserved and unsuspected roles in plant cell division. PLoS One 4, e6757 (2009). PubMed PMC

Zhang H., et al. , Role of the BUB3 protein in phragmoplast microtubule reorganization during cytokinesis. Nat. Plants 4, 485–494 (2018). PubMed

Luo Y., Ahmad E., Liu S.-T., MAD1: Kinetochore receptors and catalytic mechanisms. Front. Cell Dev. Biol. 6, 1–10 (2018). PubMed PMC

Maddox P. S., Hyndman F., Monen J., Oegema K., Desai A., Functional genomics identifies a Myb domain-containing protein family required for assembly of CENP-A chromatin. J. Cell Biol. 176, 757–763 (2007). PubMed PMC

Steiner F. A., Henikoff S., Holocentromeres are dispersed point centromeres localized at transcription factor hotspots. Elife 3, 1–22 (2014). PubMed PMC

Lermontova I., et al. , Arabidopsis KINETOCHORE NULL2 is an upstream component for centromeric histone H3 variant cenH3 deposition at centromeres. Plant Cell 25, 3389–404 (2013). PubMed PMC

Pintard L., Bowerman B., Mitotic cell division in Caenorhabditis elegans. Genetics 211, 35–73 (2019). PubMed PMC

Vondrak T., et al. , Complex sequence organization of heterochromatin in the holocentric plant Cuscuta europaea elucidated by the computational analysis of nanopore reads. Comput. Struct. Biotechnol. J. 19, 2179–2189 (2021). PubMed PMC

Dimitrova Y. N., Jenni S., Valverde R., Khin Y., Harrison S. C., Structure of the MIND complex defines a regulatory focus for yeast kinetochore assembly. Cell 167, 1014–1027.e12 (2016). PubMed PMC

Screpanti E., et al. , Direct binding of Cenp-C to the Mis12 complex joins the inner and outer kinetochore. Curr. Biol. 21, 391–398 (2011). PubMed PMC

Przewloka M. R., et al. , CENP-C is a structural platform for kinetochore assembly. Curr. Biol. 21, 399–405 (2011). PubMed

Tromer E. C., Wemyss T. A., Ludzia P., Waller R. F., Akiyoshi B., Repurposing of synaptonemal complex proteins for kinetochores in Kinetoplastida. Open Biol. 11, 210049 (2021). PubMed PMC

Butenko A., et al. , Evolution of metabolic capabilities and molecular features of diplonemids, kinetoplastids, and euglenids. BMC Biol. 18, 1–28 (2020). PubMed PMC

Karg T., Elting M. W., Vicars H., Dumont S., Sullivan W., The chromokinesin Klp3a and microtubules facilitate acentric chromosome segregation. J. Cell Biol. 216, 1597–1608 (2017). PubMed PMC

Vicars H., Karg T., Warecki B., Bast I., Sullivan W., Kinetochore-independent mechanisms of sister chromosome separation. PLOS Genet. 17, e1009304 (2021). PubMed PMC

Swentowsky K. W., et al. , Distinct kinesin motors drive two types of maize neocentromeres. Genes Dev. 34, 1239–1251 (2020). PubMed PMC

Dawe R. K., et al. , A Kinesin-14 motor activates neocentromeres to promote meiotic drive in maize. Cell 173, 839–850 (2018). PubMed

Dellaporta S. L., Wood J., Hicks J. B., A plant DNA minipreparation: Version II. Plant Mol. Biol. Rep. 1, 19–21 (1983).

Vondrak T., et al. , Characterization of repeat arrays in ultra-long nanopore reads reveals frequent origin of satellite DNA from retrotransposon-derived tandem repeats. Plant J. 101, 484–500 (2020). PubMed PMC

Zimin A. V., et al. , The MaSuRCA genome assembler. Bioinformatics 29, 2669–2677 (2013). PubMed PMC

Cheng H., Concepcion G. T., Feng X., Zhang H., Li H., Haplotype-resolved de novo assembly using phased assembly graphs with hifiasm. Nat. Methods 18, 170–175 (2021). PubMed PMC

Manni M., Berkeley M. R., Seppey M., Simão F. A., Zdobnov E. M., BUSCO update: Novel and streamlined workflows along with broader and deeper phylogenetic coverage for scoring of eukaryotic, prokaryotic, and viral genomes. Mol. Biol. Evol. 38, 4647–4654 (2021). PubMed PMC

Gurevich A., Saveliev V., Vyahhi N., Tesler G., QUAST: Quality assessment tool for genome assemblies. Bioinformatics 29, 1072–1075 (2013). PubMed PMC

Marçais G., Kingsford C., A fast, lock-free approach for efficient parallel counting of occurrences of k-mers. Bioinformatics 27, 764–770 (2011). PubMed PMC

Vurture G. W., et al. , GenomeScope: Fast reference-free genome profiling from short reads. Bioinformatics 33, 2202–2204 (2017). PubMed PMC

Grabherr M. G., et al. , Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat. Biotechnol. 29, 644–652 (2011). PubMed PMC

Dobin A., et al. , STAR: Ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013). PubMed PMC

Li H., et al. , The sequence alignment/map format and SAMtools. Bioinformatics 25, 2078–2079 (2009). PubMed PMC

Pertea M., et al. , StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat. Biotechnol. 33, 290–295 (2015). PubMed PMC

Emms D. M., Kelly S., OrthoFinder: Phylogenetic orthology inference for comparative genomics. Genome Biol. 20, 1–14 (2019). PubMed PMC

Dunn N. A., et al. , Apollo: Democratizing genome annotation. PLOS Comput. Biol. 15, e1006790 (2019). PubMed PMC

van Hooff J. J., Tromer E., van Wijk L. M., Snel B., Kops G. J., Evolutionary dynamics of the kinetochore network in eukaryotes as revealed by comparative genomics. EMBO Rep. 18, 1559–1571 (2017). PubMed PMC

Sun G., et al. , Large-scale gene losses underlie the genome evolution of parasitic plant Cuscuta australis. Nat. Commun. 9, 2683 (2018). PubMed PMC

Vogel A., et al. , Footprints of parasitism in the genome of the parasitic flowering plant Cuscuta campestris. Nat. Commun. 9, 2515 (2018). PubMed PMC

Hoshino A., et al. , Genome sequence and analysis of the Japanese morning glory Ipomoea nil. Nat. Commun. 7, 13295 (2016). PubMed PMC

Brankovics B., et al. , GRAbB: Selective assembly of genomic regions, a new niche for genomic research. PLoS Comput. Biol. 12, e1004753 (2016). PubMed PMC

Birney E., Clamp M., Durbin R., Genewise and genomewise. Genome Res. 14, 988–995 (2004). PubMed PMC

Edgar R. C., MUSCLE: Multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res. 32, 1792–1797 (2004). PubMed PMC

Novák P., Neumann P., Macas J., Global analysis of repetitive DNA from unassembled sequence reads using RepeatExplorer2. Nat. Protoc. 15, 3745–3776 (2020). PubMed

Bailey T. L., Elkan C., Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Proc. Int. Conf. Intell. Syst. Mol. Biol. 2, 28–36 (1994). PubMed

Crooks G. E., Hon G., Chandonia J.-M., Brenner S. E., WebLogo: A sequence logo generator. Genome Res. 14, 1188–1190 (2004). PubMed PMC

Weisshart K., Fuchs J., Schubert V., Structured illumination microscopy (SIM) and photoactivated localization microscopy (PALM) to analyze the abundance and distribution of RNA polymerase II molecules on flow-sorted Arabidopsis nuclei. Bio-protocol 6, 1–32 (2016). PubMed

Neumann P., Novák P., Macas J., Project PRJEB35300: Characterization of transcriptomes in Cuscuta spp. European Nucleotide Archive. https://www.ebi.ac.uk/ena/browser/view/PRJEB35300. Deposited 8 November 2019.

Macas J., Novák P., Neumann P., Project PRJEB42863: Genome sequencing of Cuscuta europaea. European Nucleotide Archive. https://www.ebi.ac.uk/ena/browser/view/PRJEB42863. Deposited 3 February 2021.

Neumann P., Novák P., Macas J., Project PRJEB51495: Investigation of changes associated with the formation of holocentric chromosomes in Cuscuta spp. European Nucleotide Archive. https://www.ebi.ac.uk/ena/browser/view/PRJEB51495. Deposited 9 March 2022.

Novák P., Neumann P., Macas J., Submitted GenBank sequence GCA_945859875.1: Genome assembly of Cuscuta europaea. NCBI Datasets. https://www.ncbi.nlm.nih.gov/data-hub/genome/GCA_945859875.1/. Deposited 27 November 2022.

Novák P., Neumann P., Macas J., Submitted GenBank sequence GCA_945859915.1: Genome assembly of Cuscuta epithymum. NCBI Datasets. https://www.ncbi.nlm.nih.gov/data-hub/genome/GCA_945859915.1/. Deposited 27 November 2022.

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