Abstract
The amino acid biosynthetic pathway of invasive pathogenic fungi has been studied as a potential antifungal drug target. Studies of the disruption of genes involved in amino acid biosynthesis have demonstrated the importance of this pathway in the virulence of Cryptococcus neoformans. Here, we identified the MET5 (CNL05500) and MET10 (CNG03990) genes in this pathway, both encoding sulfite reductase, which catalyzes the reduction of sulfite to sulfide. The MET14 (CNE03880) gene was also identified, which is responsible for the conversion of sulfate to sulfite. The use of cysteine as a sulfur source led to the production of methionine via hydrogen sulfide synthesis mediated by CYS4 (CNA06170), CYS3 (CNN01730), and MST1 (CND03690). MST1 exhibited high homology with the TUM1 gene of Saccharomyces cerevisiae, which has functional similarity with the 3-mercaptopyruvate sulfurtransferase (3-MST) gene in humans. Although the hypothesis that hydrogen sulfide is produced from cysteine via CYS4, CYS3, and MST1 warrants further study, the new insight into the metabolic pathway of sulfur-containing amino acids in C. neoformans provided here indicates the usefulness of this system in the development of screening tools for antifungal drug agents.
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The GenBank accession number for raw and processed RNA sequence data is GSE153693.
References
Altschul SF, Gish W, Miller W et al (1990) Basic local alignment search tool. J Mol Biol 215:403–410
Attarian R, Hu G, Sánchez-León E et al (2018) The monothiol glutaredoxin Grx4 regulates iron homeostasis and virulence in Cryptococcus neoformans. mBio. https://doi.org/10.1128/mBio.02377-18
Bachhawat AK, Yadav AK (2010) Metabolic pathways as drug targets: Targeting the sulphur assimilatory pathways of yeast and fungi for novel drug discovery. In: Ahmad I, Owais M, Shahid M, Aqil F (eds) Combating fungal infections. Springer, Berlin, Heidelberg, pp 327–346
Brzywczy J, Paszewski A (2002) Sulfur amino acid metabolism in Schizosaccharomyces pombe: Occurrence of two O-acetylhomoserine sulfhydrylases and the lack of the reverse transfulfuration pathway. FEMS Microbiol Lett 121:171–0174. https://doi.org/10.1111/j.1574-6968.1994.tb07095.x
Butts A, Krysan DJ (2012) Antifungal drug discovery: something old and something new. PLoS Pathog 8:9–11. https://doi.org/10.1371/journal.ppat.1002870
Chen Y, Toffaletti DL, Tenor JL et al (2014) The Cryptococcus neoformans transcriptome at the site of human meningitis. mBio 5:e01087–e1113. https://doi.org/10.1128/mBio.01087-13
Cherest H, Surdin-Kerjan Y (1992) Genetic analysis of a new mutation conferring cysteine auxotrophy in Saccharomyces cerevisiae: updating of the sulfur metabolism pathway. Genetics 130:52–58
de Melo AT, Martho KF, Roberto TN et al (2019) The regulation of the sulfur amino acid biosynthetic pathway in Cryptococcus neoformans: the relationship of Cys3, Calcineurin, and Gpp2 phosphatases. Sci Rep 9:1–19. https://doi.org/10.1038/s41598-019-48433-5
Dismukes WE (1988) Cryptococcal meningitis in patients with AIDS. J Infect Dis 157:624–628. https://doi.org/10.1093/infdis/157.4.624
Flannigan K, Wallace J (2015) Hydrogen sulfide based anti inflammatory and chemopreventive therapies: an experimental approach. Curr Pharm Des 21:3012–3022. https://doi.org/10.2174/1381612821666150514105413
Hajjeh RA, Brandt ME, Pinner RW (1995) Emergence of Cryptococcal disease: epidemiologic perspectives 100 years after its discovery. Epidemiol Rev 17:303–320. https://doi.org/10.1093/oxfordjournals.epirev.a036195
Hoffman CS, Winston F (1987) A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformaion of Escherichia coli. Gene 57:267–272. https://doi.org/10.1016/0378-1119(87)90131-4
Hopwood EMS, Ahmed D, Aitken SM (2014) A role for glutamate-333 of Saccharomyces cerevisiae cystathionine γ-lyase as a determinant of specificity. Biochim Biophys Acta 1844:465–472. https://doi.org/10.1016/j.bbapap.2013.11.012
Huang CW, Walker ME, Fedrizzi B et al (2016) The yeast TUM1 affects production of hydrogen sulfide from cysteine treatment during fermentation. FEMS Yeast Res 16:1–11. https://doi.org/10.1093/femsyr/fow100
Huang CW, Walker ME, Fedrizzi B et al (2017) Hydrogen sulfide and its roles in Saccharomyces cerevisiae in a winemaking context. FEMS Yeast Res 17:1–10. https://doi.org/10.1093/femsyr/fox058
Jakubowski H (2004) Molecular basis of homocysteine toxicity in humans. Cell Mol Life Sci 61:470–487. https://doi.org/10.1007/s00018-003-3204-7
Jiranek V, Langridge P, Henschke PA (1995) Regulation of hydrogen sulfide liberation in wine-producing Saccharomyces cerevisiae strains by assimilable nitrogen. Appl Environ Microbiol 61:461–467. https://doi.org/10.1128/aem.61.2.461-467.1995
Kaltdorf M, Srivastava M, Gupta SK et al (2016) Systematic identification of anti-fungal drug targets by a metabolic network approach. Front Mol Biosci 3:1–19. https://doi.org/10.3389/fmolb.2016.00022
Kashfi K, Olson KR (2013) Biology and therapeutic potential of hydrogen sulfide and hydrogen sulfide-releasing chimeras. Bone 85:689–703. https://doi.org/10.1038/jid.2014.371
Kim M, Kim S, Jung K, Bahn Y (2012) Targeted gene disruption in Cryptococcus neoformans using double-joint PCR with split dominant selectable markers. In: Brand AC, MacCallum DM (eds) Host-fungus interactions: Methods and protocols, 1st edn. Humana Press, United States, pp 67–84
Kim D, Langmead B, Salzberg SL (2015) HISAT: a fast spliced aligner with low memory requirements. Nat Methods 12:357–360. https://doi.org/10.1016/j.bbi.2017.04.008
Li SS, Mody CH (2010) Cryptococcus. Proc Am Thorac Soc 7:186–196. https://doi.org/10.1513/pats.200907-063AL
Lin X, Heitman J (2006) The biology of the Cryptococcus neoformans species complex. Annu Rev Microbiol 60:69–105. https://doi.org/10.1146/annurev.micro.60.080805.142102
Liu YG, Huang N (1998) Efficient amplification of insert end sequences from bacterial artificial chromosome clones by thermal asymmetric interlaced PCR. Plant Mol Biol Report 16:175–181. https://doi.org/10.1023/A:1007420918645
Martin M (2011) Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet J 17(1):10–12
Masselot M, de Robichon-Szulmajster H (1975) Methionine biosynthesis in Saccharomyces cerevisiae. Mol Gen Genet 139:121–132. https://doi.org/10.1007/BF00264692
Mikami Y, Shibuya N, Kimura Y et al (2011) Thioredoxin and dihydrolipoic acid are required for 3-mercaptopyruvate sulfurtransferase to produce hydrogen sulfide. Biochem J 439:479–485. https://doi.org/10.1042/BJ20110841
Moody BF, Calvert JW (2011) Emergent role of gasotransmitters in ischemia-reperfusion injury. Med Gas Res 1:3. https://doi.org/10.1186/2045-9912-1-3
Nazi I, Scott A, Sham A et al (2007) Role of homoserine transacetylase as a new target for antifungal agents. Antimicrob Agents Chemother 51:1731–1736. https://doi.org/10.1128/AAC.01400-06
Nielsen K, Marra RE, Hagen F et al (2005) Interaction between genetic background and the mating-type locus in Cryptococcus neoformans virulence potential. Genetics 171:975–983. https://doi.org/10.1534/genetics.105.045039
Nucci M, Marr KA (2005) Emerging fungal diseases. Clin Infect Dis 41:521–526. https://doi.org/10.1086/432060
Pascon RC, Ganous TM, Kingsbury JM et al (2004) Cryptococcus neoformans methionine synthase: expression analysis and requirement for virulence. Microbiology 150:3013–3023. https://doi.org/10.1099/mic.0.27235-0
Pertea M, Pertea GM, Antonescu CM et al (2015) StringTie enables improved reconstruction of a transcriptome from RNA-seq reads. Nat Biotechnol 33:290–295. https://doi.org/10.1016/j.physbeh.2017.03.040
Roemer T, Krysan DJ (2014) Antifungal drug development: challenges, unmet clinical needs, and new approaches. Cold Spring Harb Perspect Med 4:1–14. https://doi.org/10.1101/cshperspect.a019703
RStudioTeam (2016) RStudio: Integrated development environment for R.
Saag MS, Cloud GA, Graybill JR et al (1999) A comparison of itraconazole versus fluconazole as maintenance therapy for AIDS-associated Cryptococcal meningitis. Clin Infect Dis 28:297–298. https://doi.org/10.1086/515111
Shibuya N, Tanaka M, Yoshida M et al (2009) 3-Mercaptopyruvate sulfurtransferase produces hydrogen sulfide and bound sulfane sulfur in the brain. Antioxid Redox Signal 11:703–714. https://doi.org/10.1089/ars.2008.2253
Sohn MJ, Yoo SJ, Oh DB et al (2014) Novel cysteine-centered sulfur metabolic pathway in the thermotolerant methylotrophic yeast Hansenula polymorpha. PLoS ONE 9:1–10. https://doi.org/10.1371/journal.pone.0100725
Thomas D, Surdin-Kerjan Y (1997) Metabolism of sulfur amino acids in Saccharomyces cerevisiae. Microbiol Mol Biol Rev 61:503–532
Toffaletti DL, Rude TH, Johnston SA et al (1993) Gene transfer in Cryptococcus neoformans by use of biolistic delivery of DNA. J Bacteriol 175:1405–1411. https://doi.org/10.1128/jb.175.5.1405-1411.1993
Toh-e A, Ohkusu M, Shimizu K et al (2017) Novel biosynthetic pathway for sulfur amino acids in Cryptococcus neoformans. Curr Genet 64:681–696. https://doi.org/10.1007/s00294-017-0783-7
Trapnell C, Williams BA, Pertea G et al (2010) Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol 28:511–515. https://doi.org/10.1038/nbt.1621
Walton FJ, Idnurm A, Heitman J (2005) Novel gene functions required for melanization of the human pathogen Cryptococcus neoformans. Mol Microbiol 57:1381–1396. https://doi.org/10.1111/j.1365-2958.2005.04779.x
Warnes GR, Bolker B, Bonebakker L, et al (2016) Various R programming tools for plotting data. R Packag version 3:
Yamagata S (2007) O-acetylhomoserine sulfhydrylase of the fission yeast Schizosaccharomyces pombe: Partial purification, characterization, and its probable role in homocysteine biosynthesis. J Biochem 96:97–105. https://doi.org/10.1093/oxfordjournals.jbchem.a134980
Yang Z, Pascon RC, Alspaugh JA et al (2002) Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant. Microbiology 148:2617–2625. https://doi.org/10.1099/00221287-148-8-2617
Acknowledgements
This work was supported partly by the JSPS Grants-in-Aid for Scientific Research 16K00661 and 19K123891, and the research grant from the Institute of Fermentation, Osaka (IFO) to KS. PTN is a recipient of the Japanese Government (Monbukagakusho: MEXT) Scholoarship (G-2019-1-081).
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Nguyen, PT., Toh-e, A., Nguyen, NH. et al. Identification and characterization of a sulfite reductase gene and new insights regarding the sulfur-containing amino acid metabolism in the basidiomycetous yeast Cryptococcus neoformans. Curr Genet 67, 115–128 (2021). https://doi.org/10.1007/s00294-020-01112-9
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DOI: https://doi.org/10.1007/s00294-020-01112-9