Abstract

Eukaryotic genomes are generally organized in multiple chromosomes. Here we have created a functional single-chromosome yeast from a Saccharomyces cerevisiae haploid cell containing sixteen linear chromosomes, by successive end-to-end chromosome fusions and centromere deletions. The fusion of sixteen native linear chromosomes into a single chromosome results in marked changes to the global three-dimensional structure of the chromosome due to the loss of all centromere-associated inter-chromosomal interactions, most telomere-associated inter-chromosomal interactions and 67.4% of intra-chromosomal interactions. However, the single-chromosome and wild-type yeast cells have nearly identical transcriptome and similar phenome profiles. The giant single chromosome can support cell life, although this strain shows reduced growth across environments, competitiveness, gamete production and viability. This synthetic biology study demonstrates an approach to exploration of eukaryote evolution with respect to chromosome structure and function.

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Acknowledgements

We thank X. Gao, W. Zhao, and J. Li for technical help. This research was supported by the Strategic Priority Research Program of the Chinese Academy of Sciences (XDB19000000), the National Natural Science Foundation of China (31421061, 31770099, 31370120, 31230040), the National Key Basic Research Program of China (973 Program) (2011CBA00801, 2012CB721102), and the National Key Research and Development Program of China (2016YFA0500701).

Reviewer information

Nature thanks G. Liti, K. Wolfe and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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Affiliations

  1. Key Laboratory of Synthetic Biology, CAS Center for Excellence in Molecular Plant Sciences, Shanghai Institute of Plant Physiology and Ecology, Chinese Academy of Sciences, Shanghai, China

    • Yangyang Shao
    • , Ning Lu
    • , Chen Yang
    • , Guoping Zhao
    • , Xiaoli Xue
    •  & Zhongjun Qin
  2. University of Chinese Academy of Sciences, Beijing, China

    • Yangyang Shao
    • , Ning Lu
    • , Chen Cai
    •  & Ling-Li Zhang
  3. The State Key Laboratory of Molecular Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai, China

    • Zhenfang Wu
    • , Chen Cai
    • , Shanshan Wang
    • , Ling-Li Zhang
    •  & Jin-Qiu Zhou
  4. Frasergen Bioinformatics Co., Ltd, Wuhan, China

    • Fan Zhou
    • , Shijun Xiao
    • , Lin Liu
    •  & Xiaofei Zeng
  5. Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center at Shanghai, Shanghai, China

    • Huajun Zheng
    •  & Guoping Zhao
  6. Beijing Institute of Biotechnology, Beijing, China

    • Zhihu Zhao
  7. Department of Microbiology and Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, New Territories, Hong Kong SAR, China

    • Guoping Zhao
  8. State Key Laboratory of Genetic Engineering, Department of Microbiology, School of Life Sciences and Institute of Biomedical Sciences, Fudan University, Shanghai, China

    • Guoping Zhao

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Contributions

Z.Q. and X.X. designed and analysed all experiments. J.-Q.Z. and G.Z. contributed to the experiment designs and data evaluation. Y.S. constructed the single-chromosome yeast and conducted the scanning electron microscopy characterization. N.L. conducted the PFGE confirmation, growth characterization and cell cycle experiments. C.C. and L.-L.Z. conducted meiosis and genotoxin sensitivity experiments. Z.W. and S.W. conducted telomere characterization. S.X., X.Z. and H.Z. performed genome analysis. Z.Z. designed the Hi-C and part of the RNA-seq experiments and data interpretation. F.Z., L.L. and Z.Z. performed chromosome Hi-C data analysis. F.Z. analysed the RNA-seq data. X.X. and C.Y. analysed the phenotype microarray data. X.X. wrote the primary manuscript with contributions from J.-Q.Z., Z.Q., G.Z., Z.Z., C.Y., S.X. and Z.W.

Competing interests

The authors declare no competing interests.

Corresponding authors

Correspondence to Guoping Zhao or Jin-Qiu Zhou or Xiaoli Xue or Zhongjun Qin.

Extended data figures and tables

  1. Extended Data Fig. 1 Theoretical XhoI digestion pattern of chromosome ends.

    The X, STR, and Y′ elements in each subtelomeric regions are marked with black, grey and white boxes, respectively. The TG1–3 telomeric sequences are marked with arrow tips. The XhoI digestion sites in telomere regions are indicated, and the numbers in kb in parenthesis indicate the sizes of DNA fragments recognized by the TG1–3 probe in the Southern blot analysis.

  2. Extended Data Fig. 2 De novo sequence comparison of BY4742 (light grey) and SY14 (dark grey) genomes.

    The chromosomes are labelled with Roman numerals of the yeast reference genome. The telomeres (blue), centromeres (red) and telomere-associated repeats (green) that were cut by experimental design are shown in BY4742 chromosomes. Sequence deletions and insertions identified by genomic comparison between BY4742 and SY14 are highlighted in purple and black, respectively, in SY14 chromosomes.

  3. Extended Data Fig. 3 Comparison of the chromosomal interactions of SY6, SY13 and SY14 cells with those of BY4742 cells.

    a, Z-score difference heatmaps. Bin length, 10 kb; red and blue show increased and decreased chromatin interactions, respectively. Green box highlights the interactions of the chromosome XV centromere with other chromosomes. b, Venn diagram of the number of significant (P < 0.01, q < 0.01) ‘inter’- and intra-chromosomal interactions (referring to their locations in the BY4742 genome). c, Strong chromosomal interactions of chromosome XV centromere regions in the BY4742, SY6, SY13 and SY14 genomes. The red bars indicate the centromeres and their flanking regions of 50 MboI restriction sites. Each arc throughout the chromosome XV centromere area represents one strong interaction. In SY6, SY13 and SY14, the reserved interactions are marked with black arcs and new interactions are marked with orange arcs. The green arrowheads mark the ten residual interactions near the centromere regions found in all four strains. d, 3D structures of chromosomes XV and II in SY6, SY13 and SY14 cells compared to those in BY4742 cells. The locations of the 10 residual interactions on Chr. XV and II are marked green.

  4. Extended Data Fig. 4 3D structures of single chromosomes.

    Chromosome structures in SY6, SY13 and SY14 cells are compared to those in BY4742 cells.

  5. Extended Data Fig. 5 Directional preference plots of SY6, SY13, and SY14 cells compared to BY4742 cells.

    Red, BY4742; moss green, SY6; purple, SY13; bright green, SY14. The y-axis denotes the t-test value between the upstream and downstream interactions of each bin. A positive t-value indicates that a bin has more downstream interactions, as described previously16.

  6. Extended Data Fig. 6 Growth competition between SY14/SY14a and BY4742/BY4742a diploid cells.

    a, Blue circles represent BY4742/BY4742a (with HIS3 marker) cells that could grow only on SC-His plates; pink triangles represent SY14/SY14a (with URA3 marker) cells that could grow only on SC-Ura plates; green diamonds represent ‘fusion’ cells of BY4742/BY4742a and SY14/SY14a that could grow on both SC-His and SC-Ura plates. Data from three biological replicates are presented. b, FACS analysis of DNA content of BY4742/BY4742a and SY14/SY14a diploid cells before and after co-culture. Data are representative of two independent experiments. c, PCR verification of genomes from BY4742/BY4742a and SY14/SY14a diploid cells. H1–H3: colonies grown only on SC-His plates; HU1–3: colonies grown on both SC-His and SC-Ura plates. The BY4742/BY4742a and SY14/SY14a diploid cells before co-cultivation were used as control. Two pairs of primers, specific for genomes of BY4742/BY4742a and SY14/SY14a, were used. Data shown are representative images of two independent experiments.

  7. Extended Data Table 1 Details of the creation of a single chromosome yeast
  8. Extended Data Table 2 Information regarding long repeat sequences near chromosome ends
  9. Extended Data Table 3 SNPs and indels confirmed by re-sequencing
  10. Extended Data Table 4 Differentially expressed genes in SY14 compared to BY4742 cells

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