Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome

Shen, Yue, Wang, Yun, Chen, Tai, Gao, Feng, Gong, Jianhui, Abramczyk, Dariusz, Walker, Roy, Zhao, Hongcui, Chen, Shihong, Liu, Wei, Luo, Yisha, Müller, Carolin A., Paul-Dubois-Taine, Adrien, Alver, Bonnie, Stracquadanio, Giovanni, Mitchell, Leslie A., Luo, Zhouqing, Fan, Yanqun, Zhou, Baojin, Wen, Bo, Tan, Fengji, Wang, Yujia, Zi, Jin, Xie, Zexiong, Li, Bingzhi, Yang, Kun, Richardson, Sarah M., Jiang, Hui, French, Christopher E., Nieduszynski, Conrad A. ORCID: https://orcid.org/0000-0003-2001-076X, Koszul, Romain, Marston, Adele L., Yuan, Yingjin, Wang, Jian, Bader, Joel S., Dai, Junbiao, Boeke, Jef D., Xu, Xun, Cai, Yizhi and Yang, Huanming (2017) Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome. Science, 355 (6329). ISSN 0036-8075

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Abstract

Here, we report the successful design, construction, and characterization of a 770-kilobase synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels - including phenomics, transcriptomics, proteomics, chromosome segregation, and replication analysis - to provide a thorough and comprehensive analysis of a synthetic chromosome. Our Trans-Omics analyses reveal a modest but potentially relevant pervasive up-regulation of translational machinery observed in synII, mainly caused by the deletion of 13 transfer RNAs. By both complementation assays and SCRaMbLE (synthetic chromosome rearrangement and modification by loxP-mediated evolution), we targeted and debugged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the high-osmolarity glycerol response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain.

Item Type: Article
Additional Information: Funding Information: The project was funded by China National High Technology Research and Development Program ("863" Program: 2012AA02A708) to H.Y. and National Natural Science Foundation of China (21621004 and 21390203) to Y.Y.; a Chancellor's Fellowship from the University of Edinburgh, a start-up fund from Scottish Universities Life Sciences Alliance and Biotechnology and Biological Sciences Research Council grants (BB/M005690/1, BB/M025640/1, and BB/M00029X/1) to Y.C.; the Wellcome Trust to A.M. (090903 and 092076); U.S. NSF grants MCB-1026068 and MCB-1158201 to J.D.B.; U.S. NSF grant MCB-1445545 to J.S.B.; and U.S. Department of Energy grant DE-FG02097ER25308 to S.M.R. Work at Tsinghua University was supported by National Science Foundation of China grant 31471254 and Chinese Ministry of Science and Technology grant 2012CB725201 to J.D. Funding from ERASynBio and Agence Nationale pour la Recherche (IESY ANR-14-SYNB-0001-03) to R.K. We thank W. Zhang for many comments on the manuscript. J.D.B. and J.S.B. are founders and directors of Neochromosome Inc. J.D.B. serves as a scientific advisor to Recombinetics Inc. and Sample6, Inc. These arrangements are reviewed and managed by the committees on conflict of interest at New York University Langone Medical Center (J.D.B.) and Johns Hopkins University (J.S.B.). Y.C., H.Y., and J.D.B. conceived the concept and designed the experiments. L.M., G.S., S.M.R., J.S.B., and J.D.B. designed the synII chromosome. Y.S., Y.W., T.C.,J.G.,Y.L.,andY.C.designedexperiments.T.C.,F.G.,H.Z.,Y.S., S.C., D.A., Z.L., R.W., W.L., and F.T. performed experiments. Y.W. and J.G. analyzed genomic and RNA-seq data. Y.F., B.Z., and J.G. analyzed mass spectrometry and LC-MS data. Y.S. and D.A. analyzed synII cell cycle data. C.M. and C.N. analyzed synII replication timing profiles. G.M., H.M., and R.K. analyzed synII 3C/3D structure. The manuscript was written by Y.S., Y.W., T.C., F.G., J.D., and Y.C., with input from C.M., C.N., R.K., C.F., A.M., J.S.B, J.D.B., and H.Y. and with comments provided by all other authors. SynII genomic sequencing data and RNA-seq data have been deposited in the Sequence Read Archive (SRA) sequence database (www.ncbi.nlm.nih.gov/sra/), accession code SRP062892. Proteomics data have been submitted to the ProteomeXchange (www.proteomexchange.org/), accession code PXD003435. The metabolomics and lipidomics data have been deposited to the MetaboLights metabolomics repository (accession number MTBLS296; accessed via link www.ebi.ac.uk/metabolights/MTBLS296). The designed and sequenced synII information is uploaded with accession codes CP013607 and CP013608 on GenBank. Additional information [synII design diagram, PCRTag sequences, variants in physical strain (yeast-chr02-9-01), PCR primers, and summary of megachunks, chunks, and minichunks] related to synII can be accessed on the Sc2.0 website (www.syntheticyeast.org). Publisher Copyright: © 2017, American Association for the Advancement of Science. All rights reserved.
Uncontrolled Keywords: general ,/dk/atira/pure/subjectarea/asjc/1000
Faculty \ School: Faculty of Science > School of Biological Sciences
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Depositing User: LivePure Connector
Date Deposited: 07 Sep 2022 15:30
Last Modified: 22 Sep 2022 18:34
URI: https://ueaeprints.uea.ac.uk/id/eprint/87862
DOI: 10.1126/science.aaf4791

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