A single Ho-induced double-strand break at the MAT locus is lethal in Candida glabrata

Autoři: Laetitia Maroc aff001;  Youfang Zhou-Li aff001;  Stéphanie Boisnard aff002;  Cécile Fairhead aff001
Působiště autorů: Université Paris-Saclay, INRAE, CNRS, AgroParisTech, GQE—Le Moulon, Gif-sur-Yvette, France aff001;  Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), Gif-sur-Yvette, France aff002
Vyšlo v časopise: A single Ho-induced double-strand break at the MAT locus is lethal in Candida glabrata. PLoS Genet 16(10): e32767. doi:10.1371/journal.pgen.1008627
Kategorie: Research Article
doi: 10.1371/journal.pgen.1008627


Mating-type switching is a complex mechanism that promotes sexual reproduction in Saccharomycotina. In the model species Saccharomyces cerevisiae, mating-type switching is initiated by the Ho endonuclease that performs a site-specific double-strand break (DSB) at MAT, repaired by homologous recombination (HR) using one of the two silent mating-type loci, HMLalpha and HMRa. The reasons why all the elements of the mating-type switching system have been conserved in some Saccharomycotina, that do not show a sexual cycle nor mating-type switching, remain unknown. To gain insight on this phenomenon, we used the yeast Candida glabrata, phylogenetically close to S. cerevisiae, and for which no spontaneous and efficient mating-type switching has been observed. We have previously shown that expression of S. cerevisiae’s Ho (ScHo) gene triggers mating-type switching in C. glabrata, but this leads to massive cell death. In addition, we unexpectedly found, that not only MAT but also HML was cut in this species, suggesting the formation of multiple chromosomal DSBs upon HO induction. We now report that HMR is also cut by ScHo in wild-type strains of C. glabrata. To understand the link between mating-type switching and cell death in C. glabrata, we constructed strains mutated precisely at the Ho recognition sites. We find that even when HML and HMR are protected from the Ho-cut, introducing a DSB at MAT is sufficient to induce cell death, whereas one DSB at HML or HMR is not. We demonstrate that mating-type switching in C. glabrata can be triggered using CRISPR-Cas9, without high lethality. We also show that switching is Rad51-dependent, as in S. cerevisiae, but that donor preference is not conserved in C. glabrata. Altogether, these results suggest that a DSB at MAT can be repaired by HR in C. glabrata, but that repair is prevented by ScHo.

Klíčová slova:

Cell death – DNA repair – Genetic loci – Guide RNA – Polymerase chain reaction – Saccharomyces cerevisiae – Sexual reproduction – Yeast


1. Ni M, Feretzaki M, Sun S, Wang X, Heitman J. Sex in fungi. Annu Rev Genet. 2011;45: 405–430. doi: 10.1146/annurev-genet-110410-132536 21942368

2. Butler G. Fungal sex and pathogenesis. Clin Microbiol Rev. 2010;23: 140–159. doi: 10.1128/CMR.00053-09 20065328

3. Heitman J, Carter DA, Dyer PS, Soll DR. Sexual reproduction of human fungal pathogens. Cold Spring Harb Perspect Med. 2014;4. doi: 10.1101/cshperspect.a019281 25085958

4. Bennett RJ, Johnson AD. Completion of a parasexual cycle in Candida albicans by induced chromosome loss in tetraploid strains. EMBO J. 2003;22: 2505–2515. doi: 10.1093/emboj/cdg235 12743044

5. O’Gorman CM, Fuller HT, Dyer PS. Discovery of a sexual cycle in the opportunistic fungal pathogen Aspergillus fumigatus. Nature. 2009;457: 471–474. doi: 10.1038/nature07528 19043401

6. Hanson SJ, Wolfe KH. An Evolutionary Perspective on Yeast Mating-Type Switching. Genetics. 2017;206: 9–32. doi: 10.1534/genetics.117.202036 28476860

7. Egel R. Fission yeast mating-type switching: programmed damage and repair. DNA Repair (Amst). 2005;4: 525–536. doi: 10.1016/j.dnarep.2004.11.004 15811625

8. Maki T, Ogura N, Haber JE, Iwasaki H, Thon G. New insights into donor directionality of mating-type switching in Schizosaccharomyces pombe. PLoS Genet. 2018;14. doi: 10.1371/journal.pgen.1007424 29852001

9. Barsoum E, Martinez P, Aström SU. Alpha3, a transposable element that promotes host sexual reproduction. Genes Dev. 2010;24: 33–44. doi: 10.1101/gad.557310 20008928

10. Rajaei N, Chiruvella KK, Lin F, Åström SU. Domesticated transposase Kat1 and its fossil imprints induce sexual differentiation in yeast. Proc Natl Acad Sci U S A. 2014;111: 15491–15496. doi: 10.1073/pnas.1406027111 25313032

11. Lee C-S, Haber JE. Mating-type Gene Switching in Saccharomyces cerevisiae. Microbiol Spectr. 2015;3: MDNA3-0013–2014. doi: 10.1128/microbiolspec.MDNA3-0013-2014 26104712

12. Haber JE. Mating-type genes and MAT switching in Saccharomyces cerevisiae. Genetics. 2012;191: 33–64. doi: 10.1534/genetics.111.134577 22555442

13. Tomimatsu N, Mukherjee B, Harris JL, Boffo FL, Hardebeck MC, Potts PR, et al. DNA-damage-induced degradation of EXO1 exonuclease limits DNA end resection to ensure accurate DNA repair. J Biol Chem. 2017;292: 10779–10790. doi: 10.1074/jbc.M116.772475 28515316

14. White CI, Haber JE. Intermediates of recombination during mating type switching in Saccharomyces cerevisiae. EMBO J. 1990;9: 663–673. 2178924

15. Loo S, Rine J. Silencers and domains of generalized repression. Science. 1994;264: 1768–1771. doi: 10.1126/science.8209257 8209257

16. Herskowitz I. Life cycle of the budding yeast Saccharomyces cerevisiae. Microbiol Rev. 1988;52: 536–553. 3070323

17. Nickoloff JA, Singer JD, Heffron F. In vivo analysis of the Saccharomyces cerevisiae HO nuclease recognition site by site-directed mutagenesis. Mol Cell Biol. 1990;10: 1174–1179. doi: 10.1128/mcb.10.3.1174 2406563

18. Wu X, Haber JE. A 700 bp cis-Acting Region Controls Mating-Type Dependent Recombination Along the Entire Left Arm of Yeast Chromosome III. Cell. 1996;87: 277–285. doi: 10.1016/s0092-8674(00)81345-8 8861911

19. Connolly B, White CI, Haber JE. Physical monitoring of mating type switching in Saccharomyces cerevisiae. Molecular and Cellular Biology. 1988;8: 2342–2349. doi: 10.1128/mcb.8.6.2342 2841579

20. Dujon B, Sherman D, Fischer G, Durrens P, Casaregola S, Lafontaine I, et al. Genome evolution in yeasts. Nature. 2004;430: 35–44. doi: 10.1038/nature02579 15229592

21. Muller H, Hennequin C, Gallaud J, Dujon B, Fairhead C. The asexual yeast Candida glabrata maintains distinct a and alpha haploid mating types. Eukaryotic Cell. 2008;7: 848–858. doi: 10.1128/EC.00456-07 18375614

22. Ramírez-Zavaleta CY, Salas-Delgado GE, De Las Peñas A, Castaño I. Subtelomeric silencing of the MTL3 locus of Candida glabrata requires yKu70, yKu80, and Rif1 proteins. Eukaryotic Cell. 2010;9: 1602–1611. doi: 10.1128/EC.00129-10 20675581

23. Robledo-Márquez K, Gutiérrez-Escobedo G, Yáñez-Carrillo P, Vidal-Aguiar Y, Briones-Martín-Del-Campo M, Orta-Zavalza E, et al. Candida glabrata encodes a longer variant of the mating type (MAT) alpha2 gene in the mating type-like MTL3 locus, which can form homodimers. FEMS Yeast Res. 2016;16. doi: 10.1093/femsyr/fow082 27650705

24. Butler G, Kenny C, Fagan A, Kurischko C, Gaillardin C, Wolfe KH. Evolution of the MAT locus and its Ho endonuclease in yeast species. Proc Natl Acad Sci USA. 2004;101: 1632–1637. doi: 10.1073/pnas.0304170101 14745027

25. Carreté L, Ksiezopolska E, Pegueroles C, Gómez-Molero E, Saus E, Iraola-Guzmán S, et al. Patterns of Genomic Variation in the Opportunistic Pathogen Candida glabrata Suggest the Existence of Mating and a Secondary Association with Humans. Curr Biol. 2018;28: 15–27.e7. doi: 10.1016/j.cub.2017.11.027 29249661

26. Boisnard S, Zhou Li Y, Arnaise S, Sequeira G, Raffoux X, Enache-Angoulvant A, et al. Efficient Mating-Type Switching in Candida glabrata Induces Cell Death. PLoS ONE. 2015;10: e0140990. doi: 10.1371/journal.pone.0140990 26491872

27. Gabaldón T, Martin T, Marcet-Houben M, Durrens P, Bolotin-Fukuhara M, Lespinet O, et al. Comparative genomics of emerging pathogens in the Candida glabrata clade. BMC Genomics. 2013;14: 623. doi: 10.1186/1471-2164-14-623 24034898

28. Fidel PL, Cutright JL, Tait L, Sobel JD. A murine model of Candida glabrata vaginitis. J Infect Dis. 1996;173: 425–431. doi: 10.1093/infdis/173.2.425 8568305

29. Cormack BP, Falkow S. Efficient homologous and illegitimate recombination in the opportunistic yeast pathogen Candida glabrata. Genetics. 1999;151: 979–987. 10049916

30. Nickoloff JA, Chen EY, Heffron F. A 24-base-pair DNA sequence from the MAT locus stimulates intergenic recombination in yeast. Proc Natl Acad Sci USA. 1986;83: 7831–7835. doi: 10.1073/pnas.83.20.7831 3020559

31. Maroc L, Fairhead C. A new inducible CRISPR-Cas9 system useful for genome editing and study of double-strand break repair in Candida glabrata. Yeast. 2019. doi: 10.1002/yea.3440 31423617

32. Fairhead C, Dujon B. Consequences of unique double-stranded breaks in yeast chromosomes: death or homozygosis. Molec Gen Genet. 1993;240: 170–180. doi: 10.1007/BF00277054 8355651

33. Haber JE. DNA repair: the search for homology. Bioessays. 2018;40: e1700229. doi: 10.1002/bies.201700229 29603285

34. Xie Z-X, Mitchell LA, Liu H-M, Li B-Z, Liu D, Agmon N, et al. Rapid and Efficient CRISPR/Cas9-Based Mating-Type Switching of Saccharomyces cerevisiae. G3 (Bethesda). 2018;8: 173–183. doi: 10.1534/g3.117.300347 29150593

35. Zhou-Li Y, Boisnard S, Enache-Angoulvant A, Fairhead C. The complete sexual cycle of Nakaseomyces delphensis and the role of Ho in ploidy. Yeast. 2020. https://doi.org/10.1002/yea.3522

36. Muller H, Hennequin C, Dujon B, Fairhead C. Ascomycetes: the Candida MAT Locus: Comparing MAT in the Genomes of Hemiascomycetous Yeasts. 1st ed. Taylor JW, Kronstad JW, Heitman J, Casselton LA, editors Sex in Fungi. 1st ed. American Society for Microbiology; 2007. pp. 247–263.

37. White CI, Haber JE. Intermediates of recombination during mating type switching in Saccharomyces cerevisiae. EMBO J. 1990;9: 663–673. 2178924

38. Aylon Y, Liefshitz B, Bitan-Banin G, Kupiec M. Molecular dissection of mitotic recombination in the yeast Saccharomyces cerevisiae. Mol Cell Biol. 2003;23: 1403–1417. doi: 10.1128/mcb.23.4.1403-1417.2003 12556499

39. Richard G-F, Kerrest A, Lafontaine I, Dujon B. Comparative genomics of hemiascomycete yeasts: genes involved in DNA replication, repair, and recombination. Mol Biol Evol. 2005;22: 1011–1023. doi: 10.1093/molbev/msi083 15647519

40. Lin C-Y, Chen Y-C, Lo H-J, Chen K-W, Li S-Y. Assessment of Candida glabrata Strain Relatedness by Pulsed-Field Gel Electrophoresis and Multilocus Sequence Typing. J Clin Microbiol. 2007;45: 2452–2459. doi: 10.1128/JCM.00699-07 17553975

41. Brockert PJ, Lachke SA, Srikantha T, Pujol C, Galask R, Soll DR. Phenotypic Switching and Mating Type Switching of Candida glabrata at Sites of Colonization. Infect Immun. 2003;71: 7109–7118. doi: 10.1128/iai.71.12.7109-7118.2003 14638801

42. Belton J-M, Lajoie BR, Audibert S, Cantaloube S, Lassadi I, Goiffon I, et al. The Conformation of Yeast Chromosome III Is Mating Type Dependent and Controlled by the Recombination Enhancer. Cell Rep. 2015;13: 1855–1867. doi: 10.1016/j.celrep.2015.10.063 26655901

43. Gietz RD, Schiestl RH, Willems AR, Woods RA. Studies on the transformation of intact yeast cells by the LiAc/SS-DNA/PEG procedure. Yeast. 1995;11: 355–360. doi: 10.1002/yea.320110408 7785336

44. Zordan RE, Ren Y, Pan S-J, Rotondo G, De Las Peñas A, Iluore J, et al. Expression plasmids for use in Candida glabrata. G3 (Bethesda). 2013;3: 1675–1686. doi: 10.1534/g3.113.006908 23934995

45. Alani E, Cao L, Kleckner N. A method for gene disruption that allows repeated use of URA3 selection in the construction of multiply disrupted yeast strains. Genetics. 1987;116: 541–545. doi: 10.1534/genetics.112.541.test 3305158

46. Beyer HM, Gonschorek P, Samodelov SL, Meier M, Weber W, Zurbriggen MD. AQUA Cloning: A Versatile and Simple Enzyme-Free Cloning Approach. PLOS ONE. 2015;10: e0137652. doi: 10.1371/journal.pone.0137652 26360249

Článek vyšel v časopise

PLOS Genetics

2020 Číslo 10

Nejčtenější v tomto čísle

Tomuto tématu se dále věnují…


Zvyšte si kvalifikaci online z pohodlí domova

Nová éra v léčbě migrény
nový kurz
Autoři: MUDr. Eva Medová, MUDr. Tomáš Nežádal, Ph.D.

Imunitní trombocytopenie (ITP) u dospělých pacientů
Autoři: prof. MUDr. Tomáš Kozák, Ph.D., MBA

Pěnová skleroterapie
Autoři: MUDr. Marek Šlais

White paper - jak vidíme optimální péči o zubní náhrady
Autoři: MUDr. Jindřich Charvát, CSc.

Hemofilie - série kurzů

Všechny kurzy
Kurzy Doporučená témata Časopisy
Zapomenuté heslo

Nemáte účet?  Registrujte se

Zapomenuté heslo

Zadejte e-mailovou adresu se kterou jste vytvářel(a) účet, budou Vám na ni zaslány informace k nastavení nového hesla.


Nemáte účet?  Registrujte se

VIRTUÁLNÍ ČEKÁRNA ČR Jste praktický lékař nebo pediatr? Zapojte se! Jste praktik nebo pediatr? Zapojte se!