A genome-wide view of mutations in respiration-deficient mutants of Saccharomyces cerevisiae selected following carbon ion beam irradiation

  • Xiaopeng Guo
  • Miaomiao Zhang
  • Yue Gao
  • Guozhen Cao
  • Yang Yang
  • Dong LuEmail author
  • Wenjian LiEmail author
Genomics, transcriptomics, proteomics


Mitochondrial dysfunction in Saccharomyces cerevisiae was selected as a marker of ion penetration following carbon ion beam (CIB) irradiation. Respiration-deficient mutants were screened. Following confirmation of negligible spontaneous mutation, eight genetically stable S. cerevisiae respiration-deficient mutant strains and a control strain were resequenced with ~ 200-fold read depth. Strategies were established to identify and validate the particular mutations induced by CIB irradiation. In the nuclear genome, CIB irradiation mainly caused base substitutions and some small (< 100 bp) insertions/deletions (indels), which were widely distributed across the chromosomes. Although mitochondrial dysfunction was selected as a screening marker, variants in the nuclear genome were detected at a high frequency (10−7) relative to spontaneous mutations (10−9). The transition to transversion ratio for base substitutions was 0.746, which was less than that of spontaneous mutations. In the mitochondrial genome, there were very large deletions including substantial gene areas, resulting in extremely low read coverage. Meanwhile, every mutant possessed a distinctive mutation pattern, for both the nuclear and the mitochondrial genome. Nuclear genomes contained scanty mitochondrial respiration-related genes that were potentially affected by verified mutations, suggesting that variants in the mitochondrial genome may be the main drivers of respiratory deficiencies. Our study confirmed the previous finding that heavy ion beam (HIB) irradiation mainly induces substantial base substitutions and some small indels but also yielded some novel findings, in particular, novel structural variants in the mitochondrial genomes. These data will enhance the understanding of HIB-induced damage and mutations and aid in the HIB-based microbial mutation breeding.


Saccharomyces cerevisiae Carbon ion beam irradiation Respiration-deficient mutants Genome-wide resequencing Molecular mutation spectrum Mitochondrial genome 



The authors would like to thank the colleagues at HIRFL for providing high-quality carbon ion beam irradiation.


This work was funded by the Chinese Academy of Sciences Key Deployment Project (No. KFZD-SW-109), a Joint project of the Chinese Academy of Sciences and the Industrial Technology Research Institute (CAS-ITRI 201801) and the National Natural Science Fund of China (No. 11575259).

Compliance with ethical standards

Conflict of interest

The authors declare that there are no conflicts of interest.

Human and animal rights

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

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  1. Aokinakano M, Furusawa Y (2013) Misrepair of DNA double-strand breaks after exposure to heavy-ion beams causes a peak in the LET–RBE relationship with respect to cell killing in DT40 cells. J Radiat Res 54(6):1029–1035. CrossRefGoogle Scholar
  2. Bai H, Pekarek SD, Tichenor J, Eversman W, Buening DJ, Holbrook GR, Krefta RJ (2009) The Sequence Alignment-Map format and SAMtools. Bioinformatics 25(16):2078–2079. CrossRefGoogle Scholar
  3. Belfield EJ, Gan X, Mithani A, Brown C, Jiang C, Franklin K, Alvey E, Wibowo A, Jung M, Bailey K (2012) Genome-wide analysis of mutations in mutant lineages selected following fast-neutron irradiation mutagenesis of Arabidopsis thaliana. Genome Res 22(7):1306–1315. CrossRefGoogle Scholar
  4. Birch-Machin MA, Swalwell H (2010) How mitochondria record the effects of UV exposure and oxidative stress using human skin as a model tissue. Mutagenesis 25(2):101–107. CrossRefGoogle Scholar
  5. Boiteux S, Jinks-Robertson S (2013) DNA repair mechanisms and the bypass of DNA damage in Saccharomyces cerevisiae. Genetics 193(4):1025–1064. CrossRefGoogle Scholar
  6. Chen K, Wallis JW, McLellan MD, Larson DE, Kalicki JM, Pohl CS, McGrath SD, Wendl MC, Zhang Q, Locke DP (2009) BreakDancer: an algorithm for high-resolution mapping of genomic structural variation. Nat Methods 6(9):677–681. CrossRefGoogle Scholar
  7. Downs JA, Lowndes NF, Jackson SP (2000) A role for Saccharomyces cerevisiae histone H2A in DNA repair. Nature 408(6815):1001–1004. CrossRefGoogle Scholar
  8. Du Y, Li W, Yu L, Chen G, Liu Q, Luo S, Shu Q, Zhou L (2014) Mutagenic effects of carbon-ion irradiation on dry Arabidopsis thaliana seeds. Mutat Res-Gen Tox En 759(1):28–36. CrossRefGoogle Scholar
  9. Du Y, Luo S, Li X, Yang J, Cui T, Li W, Yu L, Feng H, Chen Y, Mu J (2017) Identification of substitutions and small insertion-deletions induced by carbon-ion beam irradiation in Arabidopsis thaliana. Front Plant Sci 8:1851. CrossRefGoogle Scholar
  10. Du Y, Luo S, Yu L, Cui T, Chen X, Yang J, Li X, Li W, Wang J, Zhou L (2018) Strategies for identification of mutations induced by carbon-ion beam irradiation in Arabidopsis thaliana by whole genome re-sequencing. Mutat Res-Fund Mol M 807:21–30. CrossRefGoogle Scholar
  11. Elahe A, Orlando TM, Léon S (2015) Biomolecular damage induced by ionizing radiation: the direct and indirect effects of low-energy electrons on DNA. Annu Rev Phys Chem 66(1):379–398. CrossRefGoogle Scholar
  12. Feng H, Yu Z, Chu PK (2006) Ion implantation of organisms. Mater Sci Eng R 54(3):49–120. CrossRefGoogle Scholar
  13. Foury F, Roganti T, Lecrenier N, Purnelle B (1998) The complete sequence of the mitochondrial genome of Saccharomyces cerevisiae. FEBS 440(3):325–331. CrossRefGoogle Scholar
  14. Foyer CH, Noctor G, Hodges M (2011) Respiration and nitrogen assimilation: targeting mitochondria-associated metabolism as a means to enhance nitrogen use efficiency. J Exp Bot 62(4):1467–1482. CrossRefGoogle Scholar
  15. Freel KC, Friedrich A, Schacherer J (2015) Mitochondrial genome evolution in yeasts: an all-encompassing view. FEMS Yeast Res 15(4):fov023. CrossRefGoogle Scholar
  16. Gdaniec Z, Ban B, Sowers LC, Fazakerley GV (1996) Methoxyamine-induced mutagenesis of nucleic acids - a proton NMR study of oligonucleotides containing N-4-methoxycytosine paired with adenine or guanine. Eur J Biochem 242(2):271–279. CrossRefGoogle Scholar
  17. Goodman MF, Hopkins RL, Lasken R, Mhaskar DN (1985) The biochemical basis of 5-bromouracil- and 2-aminopurine-induced mutagenesis. Basic Life Sci 31:409–423. Google Scholar
  18. Gualberto JM, Mileshina D, Wallet C, Niazi AK, Weberlotfi F, Dietrich A (2014) The plant mitochondrial genome: dynamics and maintenance. Biochimie 100(1):107–120. CrossRefGoogle Scholar
  19. Kazama Y, Ishii K, Hirano T, Wakana T, Yamada M, Ohbu S, Abe T (2017) Different mutational function of low- and high-linear energy transfer heavy-ion irradiation demonstrated by whole-genome resequencing of Arabidopsis mutants. Plant J 92(6):1020–1030. CrossRefGoogle Scholar
  20. Kiefer J, Egenolf R, Ikpeme S (2002) Heavy ion-induced DNA double-strand breaks in yeast. Radiat Res 157(2):141–148.[0141:hiidds];2Google Scholar
  21. Kim SR, Lee KS, Choi JH, Ha SJ, Kweon DH, Seo JH, Jin YS (2010) Repeated-batch fermentations of xylose and glucose-xylose mixtures using a respiration-deficient Saccharomyces cerevisiae engineered for xylose metabolism. J Biotechnol 150(3):404–407. CrossRefGoogle Scholar
  22. Li H, Durbin R (2009) Fast and accurate short read alignment with Burrows–Wheeler transform. Bioinformatics 25(14):1754–1760. CrossRefGoogle Scholar
  23. Li X, Wang J, Tan Z, Ma L, Lu D, Li W, Wang J (2018) Cd resistant characterization of mutant strain irradiated by carbon-ion beam. J Hazard Mater 353:1–8. CrossRefGoogle Scholar
  24. Liu XB, Gu QY, Yu XB, Luo W (2012) Enhancement of butanol tolerance and butanol yield in Clostridium acetobutylicum mutant NT642 obtained by nitrogen ion beam implantation. J Microbiol 50(6):1024–1028. CrossRefGoogle Scholar
  25. Luo S, Zhou L, Li W, Du Y, Yu L, Feng H, Mu J, Chen Y (2016) Mutagenic effects of carbon ion beam irradiations on dry Lotus japonicus seeds. NuclInstrum Meth B 383:123–128. CrossRefGoogle Scholar
  26. Lynch M, Sung W, Morris K, Coffey N, Landry CR, Dopman EB, Dickinson WJ, Okamoto K, Kulkarni S, Hartl DL (2008) A genome-wide view of the spectrum of spontaneous mutations in yeast. P Natl Acad Sci US A 105(27):9272–9277. CrossRefGoogle Scholar
  27. Mao SH, Jin GM, Wei ZQ, Xie HM, Zhang H (2006) Screening and identification of respiration deficiency mutants of yeasts (Saccharomyces cerevisiae) induced by heavy iron irradiation. J Isotopes 19(1):44–47Google Scholar
  28. Mao WJ, Liu ZZ, Zhu H, Zhu RR, Sun XY, Yao SD, Wang SL (2009) Selection of the respiration deficiency mutant yeast irritated by laser and optimization of the fermentation condition. J Rad Res Rad Proc 27(1):1–4Google Scholar
  29. Matuo Y, Nishijima S, Hase Y, Sakamoto A, Tanaka A, Shimizu K (2006) Specificity of mutations induced by carbon ions in budding yeast Saccharomyces cerevisiae. Mutat Res-Fund Mol M 602(1–2):7–13. CrossRefGoogle Scholar
  30. Naito K, Kusaba M, Shikazono N, Takano T, Tanaka A, Tanisaka T, Nishimura M (2005) Transmissible and nontransmissible mutations induced by irradiating Arabidopsis thaliana pollen with gamma-rays and carbon ions. Genetics 169(2):881–889. CrossRefGoogle Scholar
  31. Ortiz-Muñiz B, Carvajal-Zarrabal O, Aguilar B, Aguilar-Uscanga MG (2012) Improvement in ethanol production using respiratory deficient phenotype of a wild type yeast Saccharomyces cerevisiae ITV-01. Renew Energy 37(1):197–201. CrossRefGoogle Scholar
  32. Ossowski S, Schneeberger K, Lucaslledó JI, Warthmann N, Clark RM, Shaw RG, Weigel D, Lynch M (2010) The rate and molecular spectrum of spontaneous mutations in Arabidopsis thaliana. Science 327(5961):92–94. CrossRefGoogle Scholar
  33. Pitsikas P, Patapas JM, Cupples CG (2004) Mechanism of 2-aminopurine-stimulated mutagenesis in Escherichia coli. Mutat Res-Fund Mol M 550(1–2):25–32. CrossRefGoogle Scholar
  34. Porro D, Smeraldi C, Martegani E, Ranzi BM, Alberghina L (1994) Flow-cytometric determination of the respiratory activity in growing Saccharomyces cerevisiae populations. Biotechnol Prog 10(2):193–197. CrossRefGoogle Scholar
  35. Ravanat JL, Douki T (2016) UV and ionizing radiations induced DNA damage, differences and similarities. Radiat Phys Chem 128:92–102. CrossRefGoogle Scholar
  36. Reuter JA, Spacek DV, Snyder MP (2015) High-throughput sequencing technologies. Mol Cell 58(4):586–597. CrossRefGoogle Scholar
  37. Salazar AN, Gorter dVAR, Marcel VDB, Wijsman M, Pilar DLTC, Brickwedde A, Brouwers N, Daran JMG, Abeel T (2017) Nanopore sequencing enables near-complete de novo assembly of Saccharomyces cerevisiae reference strain CEN.PK113-7D. FEMS Yeast Res 17(7).
  38. Santivasi WL, Xia F (2014) Ionizing radiation-induced DNA damage, response, and repair. Antioxid Redox Signal 21(2):251–259. CrossRefGoogle Scholar
  39. Schillingtóth B, Sándor N, Kis E, Kadhim M, Sáfrány G, Hegyesi H (2011) Analysis of the common deletions in the mitochondrial DNA is a sensitive biomarker detecting direct and non-targeted cellular effects of low dose ionizing radiation. Mutat Res-Fund Mol M 716(1–2):33–39. CrossRefGoogle Scholar
  40. Seifert EL, Fiehn O, Bezaire V, Bickel DR, Wohlgemuth G, Adams SH, Harper ME (2010) Long-chain fatty acid combustion rate is associated with unique metabolite profiles in skeletal muscle mitochondria. PLoS One 5(3):e9834. CrossRefGoogle Scholar
  41. Shaughnessy DT, McAllister K, Worth L, Haugen AC, Meyer JN, Domann FE, Houten BV, Mostoslavsky R, Bultman SJ, Baccarelli AA (2014) Mitochondria, energetics, epigenetics, and cellular responses to stress. Environ Health Perspect 122(12):1271–1278. CrossRefGoogle Scholar
  42. Solieri L (2010) Mitochondrial inheritance in budding yeasts: towards an integrated understanding. Trends Microbiol 18(11):521–530. CrossRefGoogle Scholar
  43. Strope PK, Skelly DA, Kozmin SG, Mahadevan G, Stone EA, Magwene PM, Dietrich FS, McCusker JH (2015) The 100-genomes strains, an S. cerevisiae resource that illuminates its natural phenotypic and genotypic variation and emergence as an opportunistic pathogen. Genome Res 25(5):762–774. CrossRefGoogle Scholar
  44. Szumiel I (2015) Ionizing radiation-induced oxidative stress, epigenetic changes and genomic instability: the pivotal role of mitochondria. Int J Radiat Biol 91(1):1–12. CrossRefGoogle Scholar
  45. Taanman JW (1999) The mitochondrial genome: structure, transcription, translation and replication. B BA-Bioenergetics 1410(2):103–123. CrossRefGoogle Scholar
  46. Tanaka A, Shikazono N, Hase Y (2010) Studies on biological effects of ion beams on lethality, molecular nature of mutation, mutation rate, and spectrum of mutation phenotype for mutation breeding in higher plants. J Radiat Res 51(3):223–233. CrossRefGoogle Scholar
  47. Terato H, Tanaka R, Nakaarai Y, Nohara T, Doi Y, Iwai S, Hirayama R, Furusawa Y, Ide H (2008) Quantitative analysis of isolated and clustered DNA damage induced by gamma-rays, carbon ion beams, and iron ion beams. J Radiat Res 49(2):133–146. CrossRefGoogle Scholar
  48. Wallace DC (1992) Diseases of the mitochondrial DNA. Annu Rev Biochem 61(61):1175–1212. CrossRefGoogle Scholar
  49. Wang Y, Xu C, Du LQ, Cao J, Liu JX, Su X, Zhao H, Fan FY, Wang B, Katsube T (2013) Evaluation of the comet assay for assessing the dose-response relationship of DNA damage induced by ionizing radiation. Int J Mol Sci 14(11):22449–22461. CrossRefGoogle Scholar
  50. Xu A, Yao J, Yu L, Lv S, Wang J, Yan B, Yu Z (2004) Mutation of Gluconobacter oxydans and Bacillus megaterium in a two-step process of L-ascorbic acid manufacture by ion beam. J Appl Microbiol 96(6):1317–1323. CrossRefGoogle Scholar
  51. Xu TT, Bai ZZ, Wang LJ, He BF (2010) Breeding of D(-)-lactic acid high producing strain by low-energy ion implantation and preliminary analysis of related metabolism. Applied Biochem Biotech 160(2):314–321. CrossRefGoogle Scholar
  52. Zhang H, Lu D, Li X, Feng Y, Cui Q, Song X (2018a) Heavy ion mutagenesis combined with triclosan screening provides a new strategy for improving the arachidonic acid yield in Mortierella alpina. BMC Biotechnol 18(1):23. CrossRefGoogle Scholar
  53. Zhang MM, Cao GZ, Guo XP, Gao Y, Li WJ, Lu D (2018b) A comet assay for DNA damage and repair after exposure to carbon-ion beams or X-rays in Saccharomyces cerevisiae. Dose-Response 16(3):1–9. Google Scholar
  54. Zhang W, Zhao G, Luo Z, Lin Y, Wang L, Guo Y, Wang A, Jiang S, Jiang Q, Gong J (2017) Engineering the ribosomal DNA in a megabase synthetic chromosome. Science 355(6329):eaaf3981. CrossRefGoogle Scholar
  55. Zhao XT, Feng JB, Yu Wen LI, Luo Q, Yang XC, Xue LU, Chen DQ, Liu QJ (2012) Identification of two novel mitochondrial DNA deletions induced by ionizing radiation. Biomed Environ Sci 25(5):533–541. Google Scholar
  56. Zhou X, Li N, Wang Y, Wang Y, Zhang X, Zhang H (2011) Effects of X-irradiation on mitochondrial DNA damage and its supercoiling formation change. Mitochondrion 11(6):886–892. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Institute of Modern PhysicsChinese Academy of SciencesLanzhouChina
  2. 2.College of Life ScienceUniversity of Chinese Academy of SciencesBeijingChina
  3. 3.Gansu Key Laboratory of Microbial Resources Exploition and ApplicationLanzhouChina
  4. 4.Department of PharmacologySchool of Preclinical Medicine of Xinjiang Medical UniversityUrumqiChina
  5. 5.School of Life Science and EngineeringLanzhou University of TechnologyLanzhouChina

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