Telomeric ORFs (TLOs) in Candida spp. Encode Mediator Subunits That Regulate Distinct Virulence Traits

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.

Fungal Mediator Tail Subunits Contain Classical Transcriptional Activation Domains

“…Of the two TLO genes in C. dubliniensis, TLO1 is expressed at 50-fold higher levels than TLO2 under standard growth conditions and can complement most phenotypes associated in a strain with both deleted TLO genes (12). We tested the impact of the Tlo-associated TAD in C. dubliniensis by removing the C-terminal TAD in the only source of TLO1.…”

“…Complete deletion of C. dubliniensis TLO1 and TLO2 results in the inability to grow on galactose as its sole carbon source or under conditions of oxidative stress (12). These phenotypes are complemented by expressing full-length TLO1 in the mutant strain (12). To test whether the C-terminal TAD of Tlo1 was important for these adaptive responses, we tried to complement these phenotypes with the truncated Tlo1.…”

“…Because the amplification of the C. albicans TLO genes is the most striking difference from the highly syntenic C. dubliniensis genome, it has been postulated that it is an important factor in the virulence of C. albicans (10,11). Our recent finding that this amplification leads to a large population of free Tlo protein (in addition to the Mediator-associated form) in C. albicans (9) but not in C. dubliniensis (12) has led us to investigate the functional properties that the Tlo/Med2 protein alone may possess that would allow it to influence virulence.…”

Metabolic regulation in model ascomycetes – adjusting similar genomes to different lifestyles

“…This multigene family of Med2 subunits indicates increased functional potential of the C. albicans Mediator complex because the 14 TLO genes fall into at least three distinct clades [26] based on splicing patterns, expression levels, and protein localization within the cell. The closely related species C. dubliniensis lacks the TLO amplification seen in C. albicans and is considerably less virulent, suggesting that this increased potential for variety in the makeup of the Mediator complex may have an impact on cellular function [27]. Although direct assessment of the role of Med2 in C. albicans is complicated because it is encoded by a gene family, deletion of MED3 resulted in loss of the Med2 protein from the Mediator complex.…”

Candida albicans morphology: still in focus

“…Clearly, in vivo hyphal formation is mediated by a mix of environmental signals in the host, which includes hypoxia, temperature, CO 2 concentrations, and nutrient conditions, which in turn require a complex network of signaling factors and modules [33,34]. An interesting connection between filament regulation and the general transcription machinery has been found by the discovery that the mediator complex is directly linked to transcriptional regulation of hyphal regulators and hypha-associated genes [35][36][37]. Farnesol is the dominant quorum-sensing molecule and acts via different pathways.…”