1. Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China 2. Frontiers Research Institute for Synthetic Biology, Tianjin University, Tianjin 300072, China
Genomic rearrangements play a crucial role in shaping biological phenotypic diversity and driving species evolution. Synthetic chromosome rearrangement and modification by LoxP-mediated evolution (SCRaMbLE) has been applied to explore large-scale genomic rearrangements, yet it has been observed that these rearrangements occur exclusively in genomic regions containing loxPsym sites. Here, we found that SCRaMbLE of synthetic yeast harboring synthetic chromosome V and X can generate a variety of synthetic segment insertions into wild-type chromosomes, ranging from 1 to 300 kb. Furthermore, it was revealed that the novel insertions impacted the transcriptional level of neighboring regions and affected the production of exemplar pathway of zeaxanthin. Collectively, our results improve the understanding of the ability of SCRaMbLE to generate complex structural variations in nonsynthetic regions and provide a potential model to explore genomic transposable events.
J O Korbel , A E Urban , J P Affourtit , B Godwin , F Grubert , J F Simons , P M Kim , D Palejev , N J Carriero , L Du . et al.. Paired-end mapping reveals extensive structural variation in the human genome. Science, 2007, 318(5849): 420–426 https://doi.org/10.1126/science.1149504
2
M Alonge , X Wang , M Benoit , S Soyk , L Pereira , L Zhang , H Suresh , S Ramakrishnan , F Maumus , D Ciren . et al.. Major impacts of widespread structural variation on gene expression and crop improvement in tomato. Cell, 2020, 182(1): 145–161 https://doi.org/10.1016/j.cell.2020.05.021
3
J Peter , Chiara M De , A Friedrich , J X Yue , D Pflieger , A Bergström , A Sigwalt , B Barre , K Freel , A Llored . et al.. Genome evolution across 1,011 Saccharomyces cerevisiae isolates. Nature, 2018, 556(7701): 339–344 https://doi.org/10.1038/s41586-018-0030-5
4
J X Yue , J Li , L Aigrain , J Hallin , K Persson , K Oliver , A Bergström , P Coupland , J Warringer , M C Lagomarsino . et al.. Contrasting evolutionary genome dynamics between domesticated and wild yeasts. Nature Genetics, 2017, 49(6): 913–924 https://doi.org/10.1038/ng.3847
5
J Kreplak , M A Madoui , P Cápal , P Novák , K Labadie , G Aubert , P E Bayer , K K Gali , R A Syme , D Main . et al.. A reference genome for pea provides insight into legume genome evolution. Nature Genetics, 2019, 51(9): 1411–1422 https://doi.org/10.1038/s41588-019-0480-1
6
H Chen , C Li , X Peng , Z Zhou , J N Weinstein , H Liang , S J Caesar-Johnson , J A Demchok , I Felau , M Kasapi . et al.. A pan-cancer analysis of enhancer expression in nearly 9000 patient samples. Cell, 2018, 173(2): 386–399 https://doi.org/10.1016/j.cell.2018.03.027
7
G Fudenberg , G Getz , M Meyerson , L A Mirny . High order chromatin architecture shapes the landscape of chromosomal alterations in cancer. Nature Biotechnology, 2011, 29(12): 1109–1113 https://doi.org/10.1038/nbt.2049
8
Y Wu , B Z Li , M Zhao , L A Mitchell , Z X Xie , Q H Lin , X Wang , W H Xiao , Y Wang , X Zhou . et al.. Bug mapping and fitness testing of chemically synthesized chromosome X. Science, 2017, 355(6329): eaaf4706 https://doi.org/10.1126/science.aaf4706
9
Z X Xie , B Z Li , L A Mitchell , Y Wu , X Qi , Z Jin , B Jia , X Wang , B X Zeng , H M Liu . et al.. “Perfect” designer chromosome V and behavior of a ring derivative. Science, 2017, 355(6329): eaaf4704 https://doi.org/10.1126/science.aaf4704
10
S Zhou , Y Wu , Y Zhao , Z Zhang , L Jiang , L Liu , Y Zhang , J Tang , Y J Yuan . Dynamics of synthetic yeast chromosome evolution shaped by hierarchical chromatin organization. National Science Review, 2023, 10(5): nwad073 https://doi.org/10.1093/nsr/nwad073
11
H Zhang , X Fu , X Gong , Y Wang , H Zhang , Y Zhao , Y Shen . Systematic dissection of key factors governing recombination outcomes by GCE-SCRaMbLE. Nature Communications, 2022, 13(1): 5836 https://doi.org/10.1038/s41467-022-33606-0
12
B A Blount , X Lu , M R M Driessen , D Jovicevic , M I Sanchez , K Ciurkot , Y Zhao , S Lauer , R M Mckiernan , G O F Gowers . et al.. Synthetic yeast chromosome XI design provides a testbed for the study of extrachromosomal circular DNA dynamics. Cell Genomics, 2023, 3(11): 100418 https://doi.org/10.1016/j.xgen.2023.100418
13
S Zhou , Y Wu , Z X Xie , B Jia , Y J Yuan . Directed genome evolution driven by structural rearrangement techniques. Chemical Society Reviews, 2021, 50(22): 12788–12807 https://doi.org/10.1039/D1CS00722J
14
Y Zhao , C Coelho , A L Hughes , L Lazar-Stefanita , S Yang , A N Brooks , R S K Walker , W Zhang , S Lauer , C Hernandez . et al.. Debugging and consolidating multiple synthetic chromosomes reveals combinatorial genetic interactions. Cell, 2023, 186(24): 5220–5236 https://doi.org/10.1016/j.cell.2023.09.025
15
Z Gvozdenov , Z Barcutean , K Struhl . Functional analysis of a random-sequence chromosome reveals a high level and the molecular nature of transcriptional noise in yeast cells. Molecular Cell, 2023, 83(11): 1786–1797 https://doi.org/10.1016/j.molcel.2023.04.010
16
Y Xiong , H Zhang , S Zhou , L Ma , W Xiao , Y Wu , Y J Yuan . Structural variations and adaptations of synthetic chromosome ends driven by SCRaMbLE in haploid and diploid yeasts. ACS Synthetic Biology, 2023, 12(3): 689–699 https://doi.org/10.1021/acssynbio.2c00424
17
J Steensels , A Gorkovskiy , K J Verstrepen . SCRaMbLEing to understand and exploit structural variation in genomes. Nature Communications, 2018, 9(1): 1937 https://doi.org/10.1038/s41467-018-04308-3
18
Y Shen , F Gao , Y Wang , Y Wang , J Zheng , J Gong , J Zhang , Z Luo , D Schindler , Y Deng . et al.. Dissecting aneuploidy phenotypes by constructing Sc2.0 chromosome VII and SCRaMbLEing synthetic disomic yeast. Cell Genomics, 2023, 3(11): 100364 https://doi.org/10.1016/j.xgen.2023.100364
19
J Wang , Z X Xie , Y Ma , X R Chen , Y Q Huang , B He , B Jia , B Z Li , Y J Yuan . Ring synthetic chromosome V SCRaMbLE. Nature Communications, 2018, 9(1): 3783 https://doi.org/10.1038/s41467-018-06216-y
20
Y Wu , R Y Zhu , L A Mitchell , L Ma , R Liu , M Zhao , B Jia , H Xu , Y X Li , Z M Yang . et al.. In vitro DNA SCRaMbLE. Nature Communications, 2018, 9(1): 1935 https://doi.org/10.1038/s41467-018-03743-6
21
Y Zhang , T Y Chiu , J T Zhang , S J Wang , S W Wang , L Y Liu , Z Ping , Y Wang , A Chen , W W Zhang . et al.. Systematical engineering of synthetic yeast for enhanced production of lycopene. Bioengineering, 2021, 8(1): 14 https://doi.org/10.3390/bioengineering8010014
22
B Jia , J Jin , M Han , B Li , Y Yuan . Directed yeast genome evolution by controlled introduction of trans-chromosomic structural variations. Science China: Life Sciences, 2022, 65(9): 1703–1717 https://doi.org/10.1007/s11427-021-2084-1
23
L Cheng , S Zhao , T Li , S Hou , Z Luo , J Xu , W Yu , S Jiang , M Monti , D Schindler . et al.. Large-scale genomic rearrangements boost SCRaMbLE in Saccharomyces cerevisiae. Nature Communications, 2024, 15(1): 770 https://doi.org/10.1038/s41467-023-44511-5
24
K Voigt , A Gogol-Döring , C Miskey , W Chen , T Cathomen , Z Izsvák , Z Ivics . Retargeting sleeping beauty transposon insertions by engineered zinc finger DNA-binding domains. Molecular Therapy, 2012, 20(10): 1852–1862 https://doi.org/10.1038/mt.2012.126
25
H Cao , A R Hastie , D Cao , E T Lam , Y Sun , H Huang , X Liu , L Lin , W Andrews , S Chan . et al.. Rapid detection of structural variation in a human genome using nanochannel-based genome mapping technology. GigaScience, 2014, 3(1): 34 https://doi.org/10.1186/2047-217X-3-34
26
Z X Xie , L A Mitchell , H M Liu , B Z Li , D Liu , N Agmon , Y Wu , X Li , X Zhou , B Li . et al.. Rapid and efficient CRISPR/Cas9-based mating-type switching of Saccharomyces cerevisiae. G3, 2018, 8(1): 173–183 https://doi.org/10.1534/g3.117.300347
27
G Liti , D M Carter , A M Moses , J Warringer , L Parts , S A James , R P Davey , I N Roberts , A Burt , V Koufopanou . et al.. Population genomics of domestic and wild yeasts. Nature, 2009, 458(7236): 337–341 https://doi.org/10.1038/nature07743
28
D Asker . Isolation and characterization of a novel, highly selective astaxanthin-producing marine bacterium. Journal of Agricultural and Food Chemistry, 2017, 65(41): 9101–9109 https://doi.org/10.1021/acs.jafc.7b03556
29
P Wang , H Xu , H Li , H Chen , S Zhou , F Tian , B Z Li , X Bo , Y Wu , Y J Yuan . SCRaMbLEing of a synthetic yeast chromosome with clustered essential genes reveals synthetic lethal interactions. ACS Synthetic Biology, 2020, 9(5): 1181–1189 https://doi.org/10.1021/acssynbio.0c00059
30
J S Dymond , S M Richardson , C E Coombes , T Babatz , H Muller , N Annaluru , W J Blake , J W Schwerzmann , J Dai , D L Lindstrom . et al.. Synthetic chromosome arms function in yeast and generate phenotypic diversity by design. Nature, 2011, 477(7365): 471–476 https://doi.org/10.1038/nature10403
31
L A Mitchell , A Wang , G Stracquadanio , Z Kuang , X Wang , K Yang , S Richardson , J A Martin , Y Zhao , R Walker . et al.. Synthesis, debugging, and effects of synthetic chromosome consolidation: synVI and beyond. Science, 2017, 355(6329): eaaf4831 https://doi.org/10.1126/science.aaf4831
H Xu , M Han , S Zhou , B Z Li , Y Wu , Y J Yuan . Chromosome drives via CRISPR-Cas9 in yeast. Nature Communications, 2020, 11(1): 4344 https://doi.org/10.1038/s41467-020-18222-0
34
Y X Li , Y Wu , L Ma , Z Guo , W H Xiao , Y J Yuan . Loss of heterozygosity by SCRaMbLEing. Science China. Life Sciences, 2019, 62(3): 381–393 https://doi.org/10.1007/s11427-019-9504-5
35
N Ko , R Nishihama , J R Pringle . Control of 5-FOA and 5-FU resistance by Saccharomyces cerevisiae YJL055W. Yeast, 2008, 25(2): 155–160 https://doi.org/10.1002/yea.1554
36
A J Wood , T W Lo , B Zeitler , C S Pickle , E J Ralston , A H Lee , R Amora , J C Miller , E Leung , X Meng . et al.. Targeted genome editing across species using ZFNs and TALENs. Science, 2011, 333(6040): 307 https://doi.org/10.1126/science.1207773
37
T Gaj , C A Gersbach , C F Barbas . ZFN, TALEN, and CRISPR/Cas-based methods for genome engineering. Trends in Biotechnology, 2013, 31(7): 397–405 https://doi.org/10.1016/j.tibtech.2013.04.004
38
A Fleiss , S O’donnell , T Fournier , W Lu , N Agier , S Delmas , J Schacherer , G Fischer . Reshuffling yeast chromosomes with CRISPR/Cas9. PLOS Genetics, 2019, 15(8): e1008332 https://doi.org/10.1371/journal.pgen.1008332
39
T Sultana , A Zamborlini , G Cristofari , P Lesage . Integration site selection by retroviruses and transposable elements in eukaryotes. Nature Reviews. Genetics, 2017, 18(5): 292–308 https://doi.org/10.1038/nrg.2017.7
40
M Domínguez , E Dugas , M Benchouaia , B Leduque , J M Jiménez-Gómez , V Colot , L Quadrana . The impact of transposable elements on tomato diversity. Nature Communications, 2020, 11(1): 4058 https://doi.org/10.1038/s41467-020-17874-2
41
A N Brooks , A L Hughes , S Clauder-Münster , L A Mitchell , J D Boeke , L M Steinmetz . Transcriptional neighborhoods regulate transcript isoform lengths and expression levels. Science, 2022, 375(6584): 1000–1005 https://doi.org/10.1126/science.abg0162
42
A Studer , Q Zhao , J Ross-Ibarra , J Doebley . Identification of a functional transposon insertion in the maize domestication gene tb1. Nature Genetics, 2011, 43(11): 1160–1163 https://doi.org/10.1038/ng.942
43
S Soyk , Z H Lemmon , M Oved , J Fisher , K L Liberatore , S J Park , A Goren , K Jiang , A Ramos , E Van Der Knaap . et al.. Bypassing negative epistasis on yield in tomato imposed by a domestication gene. Cell, 2017, 169(6): 1142–1155 https://doi.org/10.1016/j.cell.2017.04.032
44
R Fueyo , J Judd , C Feschotte , J Wysocka . Roles of transposable elements in the regulation of mammalian transcription. Nature Reviews: Molecular Cell Biology, 2022, 23(7): 481–497 https://doi.org/10.1038/s41580-022-00457-y
45
Y Wang , M Wang , M N Djekidel , H Chen , D Liu , F W Alt , Y Zhang . eccDNAs are apoptotic products with high innate immunostimulatory activity. Nature, 2021, 599(7884): 308–314 https://doi.org/10.1038/s41586-021-04009-w
46
F Yang , W Su , O W Chung , L Tracy , L Wang , D A Ramsden , Z Z Z Zhang . Retrotransposons hijack alt-EJ for DNA replication and eccDNA biogenesis. Nature, 2023, 620(7972): 218–225 https://doi.org/10.1038/s41586-023-06327-7
47
F Guo , D N Gopaul , G D Van Duyne . Structure of Cre recombinase complexed with DNA in a site-specific recombination synapse. Nature, 1997, 389(6646): 40–46 https://doi.org/10.1038/37925
48
J L Biedler , B A Spengler . Metaphase chromosome anomaly: association with drug resistance and cell-specific products. Science, 1976, 191(4223): 185–187 https://doi.org/10.1126/science.942798
49
C Rosswog , C Bartenhagen , A Welte , Y Kahlert , N Hemstedt , W Lorenz , M Cartolano , S Ackermann , S Perner , W Vogel . et al.. Chromothripsis followed by circular recombination drives oncogene amplification in human cancer. Nature Genetics, 2021, 53(12): 1673–1685 https://doi.org/10.1038/s41588-021-00951-7
