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Error-prone, stress-induced 3′ flap–based Okazaki fragment maturation supports cell survival

Sun, Haitao and Lu, Zhaoning and Singh, Amanpreet and Zhou, Yajing and Zheng, Eric and Zhou, Mian and Wang, Jinhui and Wu, Xiwei and Hu, Zunsong and Gu, Zhaohui and Campbell, Judith L. and Zheng, Li and Shen, Binghui (2021) Error-prone, stress-induced 3′ flap–based Okazaki fragment maturation supports cell survival. Science, 374 (6572). pp. 1252-1258. ISSN 0036-8075. PMCID PMC8852821. doi:10.1126/science.abj1013.

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How cells with DNA replication defects acquire mutations that allow them to escape apoptosis under environmental stress is a long-standing question. Here, we report that an error-prone Okazaki fragment maturation (OFM) pathway is activated at restrictive temperatures in rad27Δ yeast cells. Restrictive temperature stress activated Dun1, facilitating transformation of unprocessed 5′ flaps into 3′ flaps, which were removed by 3′ nucleases, including DNA polymerase δ (Polδ). However, at certain regions, 3′ flaps formed secondary structures that facilitated 3′ end extension rather than degradation, producing alternative duplications with short spacer sequences, such as pol3 internal tandem duplications. Consequently, little 5′ flap was formed, suppressing rad27Δ-induced lethality at restrictive temperatures. We define a stress-induced, error-prone OFM pathway that generates mutations that counteract replication defects and drive cellular evolution and survival.

Item Type:Article
Related URLs:
URLURL TypeDescription CentralArticle
Sun, Haitao0000-0001-7354-3960
Zheng, Eric0000-0003-0897-1218
Wu, Xiwei0000-0002-7071-1671
Hu, Zunsong0000-0003-3167-7148
Gu, Zhaohui0000-0003-1581-1327
Zheng, Li0000-0002-3744-185X
Shen, Binghui0000-0002-4408-407X
Alternate Title:Error-prone, stress-induced 3’ flap-based Okazaki fragment maturation supports cell survival
Additional Information:© 2021 American Association for the Advancement of Science. Received 20 April 2021; accepted 14 October 2021. We thank R. Kolodner for the yeast strains RDKY2672, RDKY2608, and RDKY2669; P. M. J. Burgers for the plasmids pBL335 (GST-Pol3), pBL338 (GAL1-Pol31), pBL340 (GAL10-Pol32), and pBL341 (Pol31/Pol32); L. Prakash and S. Prakash for the protease-deficient yeast strain YRP654 and the plasmids pBJ1445 (Flag-Pol3) and pBJ1524 (GST-Pol31/Pol32) to express the yeast recombinant Polδ complex (Pol3, Pol31, and Pol32); W.-D. Heyer for the anti-Dun1 antibody; and M. S. Wold for purified recombinant yeast replication protein A (RPA) complex. We thank H. Lou’s laboratory members and H. Dai, D. Duenas, and M. E. Budd for technical assistance in mouse and yeast genetic experiments and stimulating discussions. We thank K. Walker and S. Wilkinson for critical reading and editing of the manuscript. This work was supported by NIH grants R50 CA211397 to L.Z. and R01 CA073764 and R01 CA085344 to B.S. Research reported in this publication includes work performed by City of Hope shared resources supported by the National Cancer Institute of the NIH under award number P30 CA033572. Author contributions: H.S., Z.L., A.S., Y.Z., and M.Z. conducted yeast genetic and biochemical experiments. E.Z., J.W., X.W., Z.H., and Z.G. conducted RNA sequencing (RNA-seq), WES, and WGS and performed data analysis. J.L.C. designed yeast genetic experiments and conducted data analysis. L.Z. conducted biochemical experiments, RNA-seq, and WGS data analysis; designed and coordinated most of the experiments; and wrote the first draft of the manuscript. B.S. supervised the entire project; designed and coordinated most of the experiments; and provided input into and finalized the manuscript. The authors declare no conflicts of interest in this study. Data and materials availability: All data are available in the manuscript or the supplementary materials. Gene Expression Omnibus accession numbers for the mouse and yeast genomics datasets are GSE181154 and GSE178876, respectively.
Funding AgencyGrant Number
NIHR50 CA211397
NIHR01 CA073764
NIHR01 CA085344
NIHP30 CA033572
Issue or Number:6572
PubMed Central ID:PMC8852821
Record Number:CaltechAUTHORS:20211203-172529774
Persistent URL:
Official Citation:Error-prone, stress-induced 3' flap–based Okazaki fragment maturation supports cell survival. Haitao Sun, Zhaoning Lu, Amanpreet Singh, Yajing Zhou, Eric Zheng, Mian Zhou, Jinhui Wang, Xiwei Wu, Zunsong Hu, Zhaohui Gu, Judith L. Campbell, Li Zheng, Binghui Shen. Science, 374 (6572); DOI: 10.1126/science.abj1013
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:112188
Deposited By: Tony Diaz
Deposited On:03 Dec 2021 17:59
Last Modified:31 May 2022 18:36

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