Titrating gene function in the human fungal pathogen Candida albicans through polyadenosine tract insertion

Helene Tournu, Arielle Butts, Glen Palmer

Research output: Contribution to journalArticle

Abstract

A recent study demonstrated that the insertion of poly-adenosine (poly-A) tracts into an open reading frame can suppress expression of the encoded protein in both prokaryotic and eukaryotic species. Furthermore, the degree of suppression is proportional to the length of the poly-A insertion, which can therefore provide a reliable and predictable means to titrate a specific protein's expression. The goal of this study was to determine if this methodology can be applied to modulate the expression of proteins in the prevalent human fungal pathogen, Candida albicans. Insertion of increasing numbers of AAA codons encoding lysine at the N terminus of the C. albicans lanosterol demethylase (Erg11p) progressively diminished expression without significantly reducing the levels of mRNA. This suggests that Erg11p expression was attenuated at the posttranscriptional level. A direct correlation between the number of AAA codons inserted and C. albicans susceptibility to the Erg11p inhibitor fluconazole was also noted, indicating a progressive loss of Erg11p activity. Finally, we constructed a series of C. albicans strains with 3 to 12 AAA codons inserted at the 5' end of the ARO1 gene, which encodes a pentafunctional enzyme catalyzing five sequential steps of the aromatic amino acid biosynthetic pathway. Increasing numbers of AAA codons progressively reduced the growth rate of C. albicans in standard laboratory medium, indicating a progressive loss of ARO biosynthetic activity. These data unequivocally demonstrate the potential utility of the poly-A insertion method to examine the phenotypic consequences of titrating target protein function in C. albicans.

Original languageEnglish (US)
Article numbere00192-19
JournalmSphere
Volume4
Issue number3
DOIs
StatePublished - May 1 2019

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Candida albicans
Codon
Adenosine
Genes
Proteins
Lanosterol
Aromatic Amino Acids
Fluconazole
Biosynthetic Pathways
Open Reading Frames
Lysine
polyadenosine
Messenger RNA
Enzymes
Growth

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

Cite this

Titrating gene function in the human fungal pathogen Candida albicans through polyadenosine tract insertion. / Tournu, Helene; Butts, Arielle; Palmer, Glen.

In: mSphere, Vol. 4, No. 3, e00192-19, 01.05.2019.

Research output: Contribution to journalArticle

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