Skip to main content

Advertisement

SpringerLink
Log in

RNA-Mediated Gene Silencing in Candida albicans: Inhibition of Hyphae Formation by Use of RNAi Technology

  • Published:
Mycopathologia Aims and scope Submit manuscript

Abstract

The introduction of RNA silencing machinery in fungi has led to the promising application of RNAi methodology to knock down essential vital factor or virulence factor genes in the microorganisms. Efg1p is required for development of a true hyphal growth form which is known to be essential for interactions with human host cells and for the yeast’s pathogenesis. In this paper, we describe the development of a system for presenting and studying the RNAi function on the EFG1 gene in C. albicans. The 19-nucleotide siRNA was designed on the basis of the cDNA sequence of the EFG1 gene in C. albicans and transfection was performed by use of a modified-PEG/LiAc method. To investigate EFG1 gene silencing in siRNA-treated cells, the yeasts were grown in human serum; to induce germ tubes a solid medium was used with the serum. Quantitative changes in expression of the EFG1 gene were analyzed by measuring the cognate EFG1 mRNA level by use of a quantitative real-time RT-PCR assay. Compared with the positive control, true hyphae formation was significantly reduced by siRNA at concentrations of 1 μM, 500 nM, and 100 nM (P < 0.05). In addition, siRNA at a concentration of 1 μM was revealed to inhibit expression of the EFG1 gene effectively (P < 0.05). On the basis of the potential of post-transcriptional gene silencing to control the expression of specific genes, these techniques may be regarded as promising means of drug discovery, with applications in biomedicine and functional genomics analysis.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Gow NA, Brown AJ, Odds FC. Fungal morphogenesis and host invasion. Curr Opin Microbiol. 2002;5(4):366–71.

    Article  PubMed  CAS  Google Scholar 

  2. Rooney PJ, Klein BS. Linking fungal morphogenesis with virulence. Cell Microbiol. 2002;4(3):127–37.

    Article  PubMed  CAS  Google Scholar 

  3. Odds FC. Candida and candidosis: a review and bibliography. London: Baillière Tindall; 1988.

    Google Scholar 

  4. Phan QT, Belanger PH, Filler SG. Role of hyphal formation in interactions of Candida albicans with endothelial cells. Infect Immun. 2000;68(6):3485–90.

    Article  PubMed  CAS  Google Scholar 

  5. Srikantha T, Tsai LK, Daniels K, Soll DR. EFG1 null mutants of Candida albicans switch but cannot express the complete phenotype of white-phase budding cells. J Bacteriol. 2000;182(6):1580–91.

    Article  PubMed  CAS  Google Scholar 

  6. Felk A, Kretschmar M, Albrecht A, Schaller M, Beinhauer S, Nichterlein T, et al. Candida albicans hyphal formation and the expression of the Efg1-regulated proteinases Sap4 to Sap6 are required for the invasion of parenchymal organs. Infect Immun. 2002;70(7):3689–700.

    Article  PubMed  CAS  Google Scholar 

  7. Ernst JF. Transcription factors in Candida albicans—environmental control of morphogenesis. Microbiology. 2000;146(8):1763.

    PubMed  CAS  Google Scholar 

  8. Brown AJP, Gow NAR. Regulatory networks controlling Candida albicans morphogenesis. Trends Microbiol. 1999;7(8):333–8.

    Article  PubMed  CAS  Google Scholar 

  9. Stoldt VR, Sonneborn A, Leuker CE, Ernst JF. Efg1p, an essential regulator of morphogenesis of the human pathogen Candida albicans, is a member of a conserved class of bHLH proteins regulating morphogenetic processes in fungi. EMBO J. 1997;16(8):1982–91.

    Article  PubMed  CAS  Google Scholar 

  10. Lo HJ, Kohler JR, DiDomenico B, Loebenberg D, Cacciapuoti A, Fink GR. Nonfilamentous C. albicans mutants are avirulent. Cell. 1997;90(5):939–49.

    Article  PubMed  CAS  Google Scholar 

  11. Noffz CS, Liedschulte V, Lengeler K, Ernst JF. Functional mapping of the Candida albicans Efg1 regulator. Eukaryot Cell. 2008;7(5):881–93.

    Article  PubMed  CAS  Google Scholar 

  12. Kumamoto CA, Vinces MD. Contributions of hyphae and hypha-co-regulated genes to Candida albicans virulence. Cell Microbiol. 2005;7(11):1546–54.

    Article  PubMed  CAS  Google Scholar 

  13. Sonneborn A, Bockmuhl DP, Ernst JF. Chlamydospore formation in Candida albicans requires the Efg1p morphogenetic regulator. Infect Immun. 1999;67(10):5514–7.

    PubMed  CAS  Google Scholar 

  14. Staib P, Kretschmar M, Nichterlein T, Hof H, Morschhauser J. Transcriptional regulators Cph1p and Efg1p mediate activation of the Candida albicans virulence gene SAP5 during infection. Infect Immun. 2002;70(2):921–7.

    Article  PubMed  CAS  Google Scholar 

  15. Korting HC, Hube B, Oberbauer S, Januschke E, Hamm G, Albrecht A, et al. Reduced expression of the hyphal-independent Candida albicans proteinase genes SAP1 and SAP3 in the efg1 mutant is associated with attenuated virulence during infection of oral epithelium. J Med Microbiol. 2003;52(Pt 8):623–32.

    Article  PubMed  CAS  Google Scholar 

  16. Hoyer LL, Payne TL, Bell M, Myers AM, Scherer S. Candida albicans ALS3 and insights into the nature of the ALS gene family. Curr Genet. 1998;33(6):451–9.

    Article  PubMed  CAS  Google Scholar 

  17. Sharkey LL, McNemar MD, Saporito-Irwin SM, Sypherd PS, Fonzi WA. HWP1 functions in the morphological development of Candida albicans downstream of EFG1, TUP1, and RBF1. J Bacteriol. 1999;181(17):5273–9.

    PubMed  CAS  Google Scholar 

  18. Fu Y, Ibrahim AS, Sheppard DC, Chen YC, French SW, Cutler JE, et al. Candida albicans Als1p: an adhesin that is a downstream effector of the EFG1 filamentation pathway. Mol Microbiol. 2002;44(1):61–72.

    Article  PubMed  CAS  Google Scholar 

  19. Nakayashiki H. RNA silencing in fungi: mechanisms and applications. FEBS Lett. 2005;579(26):5950–7.

    Article  PubMed  CAS  Google Scholar 

  20. Drinnenberg IA, Weinberg DE, Xie KT, Mower JP, Wolfe KH, Fink GR, et al. RNAi in budding yeast. Science. 2009;326(5952):544–50.

    Article  PubMed  CAS  Google Scholar 

  21. Harrison BR, Yazgan O, Krebs JE. Life without RNAi: noncoding RNAs and their functions in Saccharomyces cerevisiae. Biochem Cell Biol. 2009;87(5):767–79.

    Article  PubMed  CAS  Google Scholar 

  22. Rao DD, Vorhies JS, Senzer N, Nemunaitis J. siRNA vs. shRNA: similarities and differences. Adv Drug Deliv Rev. 2009;61(9):746–59.

    Article  PubMed  CAS  Google Scholar 

  23. Jinek M, Doudna JA. A three-dimensional view of the molecular machinery of RNA interference. Nature. 2009;457(7228):405–12.

    Article  PubMed  CAS  Google Scholar 

  24. Romano N, Macino G. Quelling: transient inactivation of gene expression in Neurospora crassa by transformation with homologous sequences. Mol Microbiol. 1992;6(22):3343–53.

    Article  PubMed  CAS  Google Scholar 

  25. Rappleye CA, Engle JT, Goldman WE. RNA interference in Histoplasma capsulatum demonstrates a role for alpha-(1,3)-glucan in virulence. Mol Microbiol. 2004;53(1):153–65.

    Article  PubMed  CAS  Google Scholar 

  26. Nakayashiki H, Nguyen QB. RNA interference: roles in fungal biology. Curr Opin Microbiol. 2008;11(6):494–502.

    Article  PubMed  CAS  Google Scholar 

  27. Zhao W, Fanning ML, Lane T. Efficient RNAi-based gene family knockdown via set cover optimization. Artif Intell Med. 2005;35(1–2):61–73.

    Article  PubMed  Google Scholar 

  28. Liu H, Cottrell TR, Pierini LM, Goldman WE, Doering TL. RNA interference in the pathogenic fungus Cryptococcus neoformans. Genetics. 2002;160(2):463–70.

    PubMed  CAS  Google Scholar 

  29. Disney MD, Haidaris CG, Turner DH. Uptake and antifungal activity of oligonucleotides in Candida albicans. Proc Natl Acad Sci USA. 2003;100(4):1530–4.

    Article  PubMed  CAS  Google Scholar 

  30. Janbon G, Maeng S, Yang DH, Ko YJ, Jung KW, Moyrand F, et al. Characterizing the role of RNA silencing components in Cryptococcus neoformans. Fungal Genet Biol. 2010;47(12):1070–80.

    Article  PubMed  CAS  Google Scholar 

  31. Khatri M, Rajam MV. Targeting polyamines of Aspergillus nidulans by siRNA specific to fungal ornithine decarboxylase gene. Med Mycol. 2007;45(3):211–20.

    Article  PubMed  CAS  Google Scholar 

  32. Staab JF, White TC, Marr KA. Hairpin dsRNA does not trigger RNA interference in Candida albicans cells. Yeast. 2011;28(1):1–8.

    Article  PubMed  CAS  Google Scholar 

  33. De Backer MD, Nelissen B, Logghe M, Viaene J, Loonen I, Vandoninck S, et al. An antisense-based functional genomics approach for identification of genes critical for growth of Candida albicans. Nat Biotechnol. 2001;19(3):235–41.

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

The authors thank the Central Research Laboratory of the School of Public Health for providing laboratory facilities. This research was financially supported by a Tehran University of Medical Sciences (TUMS) grant.

Conflict of interest

The authors declare that there is no conflict of interests.

Author information

Authors and Affiliations

  1. Division of Molecular Biology, Department of Medical Mycology and Parasitology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran

    Maryam Moazeni, Parivash Kordbacheh & Sassan Rezaie

  2. Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, P.O. Box. 14155-6446, Tehran, Iran

    Mohammad Reza Khoramizadeh & Sassan Rezaie

  3. Molecular Medicine Department, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran

    Ladan Teimoori-Toolabi

  4. Department of Pharmaceutical Biotechnology, School of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran

    Zargham Sepehrizadeh

  5. Department of Biology, Islamic Azad University, Varamin-Pishva Branch, Varamin, Iran

    Fatemeh Noorbakhsh

  6. Epidemiology and Biostatistics Department, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran

    Hojat Zeraati

Authors
  1. Maryam Moazeni
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mohammad Reza Khoramizadeh
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Parivash Kordbacheh
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Zargham Sepehrizadeh
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Hojat Zeraati
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Fatemeh Noorbakhsh
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ladan Teimoori-Toolabi
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Sassan Rezaie
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Sassan Rezaie.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Moazeni, M., Khoramizadeh, M.R., Kordbacheh, P. et al. RNA-Mediated Gene Silencing in Candida albicans: Inhibition of Hyphae Formation by Use of RNAi Technology. Mycopathologia 174, 177–185 (2012). https://doi.org/10.1007/s11046-012-9539-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11046-012-9539-6

Keywords

Search

Navigation