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RNA silencing in the dermatophyte Microsporum canis

Sandy Vermout1, Jérémy Tabart, Aline Baldo

  • 1Department of Infectious & Parasitic Diseases, Parasitology, Faculty of Veterinary Medicine, University of Liège, Liège, Belgium.

FEMS Microbiology Letters
|August 8, 2007
PubMed

Insights

RNA silencing effectively down-regulated key proteases in Microsporum canis, a fungus causing animal and human dermatomycoses. This method offers a valuable tool for studying fungal virulence factors.

Area of Science:

  • Mycology
  • Molecular Biology
  • Genetics

Background:

  • Dermatomycoses, fungal skin infections, are common in pets and transmissible to humans.
  • Microsporum canis secretes proteases identified as potential virulence factors.
  • Investigating protease roles requires creating gene-deficient fungal strains, which is technically challenging.

Purpose of the Study:

  • To evaluate RNA-mediated silencing as a reverse genetic tool in dermatophytes.
  • To assess the utility of RNA silencing for down-regulating specific protease genes in Microsporum canis.

Main Methods:

  • Developed plasmid constructs to express RNA hairpins targeting SUB3 (subtilisin) and DPPIV (dipeptidyl peptidase) genes.
  • Assessed gene down-regulation by measuring protease enzymatic activity in culture supernatants.
  • Quantified gene expression levels using real-time reverse transcriptase-polymerase chain reaction (RT-PCR).

Main Results:

  • Both SUB3 and DPPIV genes were successfully down-regulated in Microsporum canis transformants.
  • Enzymatic activities and expression levels ranged from <5% to 100% of control levels.
  • RNA silencing was generally more effective for inhibiting SUB3 than DPPIV.

Conclusions:

  • RNA-mediated gene silencing is a viable functional genomics approach for Microsporum canis.
  • This technique circumvents technical difficulties associated with conventional gene disruption methods in dermatophytes.
  • RNA silencing facilitates the study of fungal virulence factors like proteases in Microsporum canis.

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