Telomeric ORFs (TLOs) in Candida spp. Encode mediator subunits that regulate distinct virulence traits.

John Haran,Tim Yeomans,Michael Church,Zhongle Liu,Hannah Boyle,Derek J Sullivan,Matthew Z Anderson,Judith Berman,Lawrence C Myers,Karsten Hokamp,Alastair B Fleming,Gary P Moran,Geraldine Butler

doi:10.1371/journal.pgen.1004658

Abstract

The TLO genes are a family of telomere-associated ORFs in the fungal pathogens Candida albicans and C. dubliniensis that encode a subunit of the Mediator complex with homology to Med2. The more virulent pathogen C. albicans has 15 copies of the gene whereas the less pathogenic species C. dubliniensis has only two (CdTLO1 and CdTLO2). In this study we used C. dubliniensis as a model to investigate the role of TLO genes in regulating virulence and also to determine whether TLO paralogs have evolved to regulate distinct functions. A C. dubliniensis tlo1Δ/tlo2Δ mutant is unable to form true hyphae, has longer doubling times in galactose broth, is more susceptible to oxidative stress and forms increased levels of biofilm. Transcript profiling of the tlo1Δ/tlo2Δ mutant revealed increased expression of starvation responses in rich medium and retarded expression of hypha-induced transcripts in serum. ChIP studies indicated that Tlo1 binds to many ORFs including genes that exhibit high and low expression levels under the conditions analyzed. The altered expression of these genes in the tlo1Δ/tlo2Δ null mutant indicates roles for Tlo proteins in transcriptional activation and repression. Complementation of the tlo1Δ/tlo2Δ mutant with TLO1, but not TLO2, restored wild-type filamentous growth, whereas only TLO2 fully suppressed biofilm growth. Complementation with TLO1 also had a greater effect on doubling times in galactose broth. The different abilities of TLO1 and TLO2 to restore wild-type functions was supported by transcript profiling studies that showed that only TLO1 restored expression of hypha-specific genes (UME6, SOD5) and galactose utilisation genes (GAL1 and GAL10), whereas TLO2 restored repression of starvation-induced gene transcription. Thus, Tlo/Med2 paralogs encoding Mediator subunits regulate different virulence properties in Candida spp. and their expansion may account for the increased adaptability of C. albicans relative to other Candida species.

Highlights

Candida albicans is a commensal yeast commonly recovered from mucosal surfaces in humans
In this study we show that a C. dubliniensis mutant missing both TLO1 and TLO2 is defective in virulence functions, including hyphal growth and stress responses but forms increased levels of biofilm
Tlo1 protein was found to interact with genes and this was associated with both gene activation and repression

Summary

Introduction

Candida albicans is a commensal yeast commonly recovered from mucosal surfaces in humans. Transcriptional profiling of C. albicans using microarray and RNA-seq technologies has revealed that rapid adaptation to local environmental conditions involves elaborate, programmed shifts in transcription pattern. Most genes transcribed by RNA polymerase II (PolII) require a carefully orchestrated series of events to initiate transcription. The formation of the preinitiation complex and recruitment of PolII requires the function of Mediator, a large multi-subunit protein complex [4,5]. Mediator is thought to be required to bridge DNA-bound transcription factors with the rest of the transcriptional machinery [6,7]. The tail region includes Med, Med and Med15/Gal and is thought to be the part of Mediator that

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