Lim Wen Jun, Lee Brenda, Farrington Zahra, Abu Alhaija Abed Alkarem, Fleming Alastair B, Sullivan Derek J, Moran Gary P
Division of Oral Biosciences, Dublin Dental University Hospital, University of Dublin, Trinity College Dublin, Dublin, Ireland.
Department of Microbiology, School of Genetics and Microbiology, University of Dublin, Trinity College Dublin, Dublin, Ireland.
Microbiology (Reading). 2025 Sep;171(9). doi: 10.1099/mic.0.001594.
is an opportunistic fungal pathogen associated with superficial and systemic infections in humans. Azole antifungal resistance in is of clinical concern, and both oral and systemic infections can be difficult to treat due to the lack of alternative antifungal drugs. Expression of a hyperactive form of the transcription factor Tac1 is a major contributor to azole resistance in isolates resulting in the increased expression of the azole efflux pump Cdr1. In this study, we investigated whether the Mediator tail component Med2, encoded by the expanded (n=14) gene family of , was required for Tac1 activity. A homozygous gain-of-function point mutation was introduced into WT, Δ and Δ strains of which enables them to express hyperactive Tac1. qRT-PCR analysis revealed that Δ- had reduced basal and fluphenazine-induced expression relative to WT- strains and exhibited reduced levels of resistance to fluconazole and terbinafine. Individual copies of representatives from each of the alpha, beta and gamma clades were reintroduced into Δ- to investigate their ability to restore Tac1-activated resistance. These studies show that alpha and beta genes could restore fluconazole resistance in the Δ- background, whereas gamma clade genes did not result in any detectable phenotypic complementation. Transcript profiling showed that reintroduction of α led to increased expression of -activated genes such as . Further analysis using ChIP-qPCR revealed that Tloα1 localizes to the drug response element which is the site where Tac1 binds to the promoter. These data have identified that the gene family is required for the expression of Tac1-mediated fluconazole resistance. However, this effect is confined to members of the alpha and beta, but not the gamma, clades.
是一种与人类浅表和全身感染相关的机会性真菌病原体。其对唑类抗真菌药的耐药性是临床关注的问题，由于缺乏替代抗真菌药物，口服和全身感染都可能难以治疗。转录因子Tac1的高活性形式的表达是导致分离株对唑类耐药的主要因素，导致唑类外排泵Cdr1的表达增加。在本研究中，我们调查了由扩展的（n = 14）基因家族编码的中介体尾部成分Med2是否是Tac1活性所必需的。将纯合的功能获得性点突变引入的野生型、Δ和Δ菌株中，使其能够表达高活性的Tac1。qRT-PCR分析显示，相对于野生型菌株，Δ-的基础和氟奋乃静诱导的表达降低，并且对氟康唑和特比萘芬的耐药水平降低。将来自α、β和γ进化枝的每个代表的单个拷贝重新引入Δ-中，以研究它们恢复Tac1激活的耐药性的能力。这些研究表明，α和β基因可以在Δ-背景中恢复氟康唑耐药性，而γ进化枝基因没有导致任何可检测到的表型互补。转录谱分析表明，α的重新引入导致了如等Tac1激活基因的表达增加。使用ChIP-qPCR的进一步分析表明，Tloα1定位于药物反应元件，该元件是Tac1与启动子结合的位点。这些数据表明基因家族是Tac1介导的氟康唑耐药性表达所必需的。然而，这种效应仅限于α和β进化枝的成员，而不是γ进化枝。
