TOP2 gene disruption reduces drug susceptibility by increasing intracellular ergosterol biosynthesis in Candida albicans.
In this study the role of the TOP2 gene in fungal drug susceptibility was investigated by disrupting and overexpressing the gene in Candida albicans. MIC determination and a spot assay showed that a top2Delta/Delta null mutant (strain T2bc) was more resistant to the antifungals tested than the wild-type (strain CAI4). Real-time RT-PCR and rhodamine 6G efflux examination showed that TOP2 did not influence the activity of drug efflux pumps. Sterol analysis with GC/high-resolution MS indicated that the intracellular ergosterol composition of the top2Delta/Delta mutant was significantly increased. Subsequently, fluorescence polarization measurements also revealed that Top2-deprived cells displayed a decrease in membrane fluidity, resulting in enhanced passive diffusion of the drugs. Quantitative real-time RT-PCR analysis further confirmed that the ERG11 gene, an essential gene in ergosterol biosynthesis, was upregulated. These results demonstrate a close relationship between the TOP2 gene and drug susceptibility in C. albicans.
Zheng
Hao
H
School of Pharmacy, Second Military Medical University, Shanghai, PR China.|Department of Pharmacology, China Pharmaceutical University, Nanjing, PR China.
Jiang
Yuan-Ying
YY
School of Pharmacy, Second Military Medical University, Shanghai, PR China.
Wang
Yan
Y
School of Pharmacy, Second Military Medical University, Shanghai, PR China.
Jia
Xin-Ming
XM
School of Pharmacy, Second Military Medical University, Shanghai, PR China.
Yan
Tian-Hua
TH
Department of Pharmacology, China Pharmaceutical University, Nanjing, PR China.
Gao
Ping-Hui
PH
School of Pharmacy, Second Military Medical University, Shanghai, PR China.
Yan
Lan
L
School of Pharmacy, Second Military Medical University, Shanghai, PR China.
Jiang
Ling-Huo
LH
School of Pharmaceutical Science and Technology, Tianjin University, PR China.
Ji
Hui
H
Department of Pharmacology, China Pharmaceutical University, Nanjing, PR China.
Cao
Yong-Bing
YB
School of Pharmacy, Second Military Medical University, Shanghai, PR China.
D000935:Antifungal Agents / Q000494:pharmacology*; D002176:Candida albicans / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D004250:DNA Topoisomerases, Type II / Q000235:genetics* / Q000378:metabolism; D025141:Drug Resistance, Fungal / Q000235:genetics; D004875:Ergosterol / Q000096:biosynthesis* / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal / Q000502:physiology*; D008826:Microbial Sensitivity Tests; D009154:Mutation
D000935:Antifungal Agents; D005656:Fungal Proteins; D004250:DNA Topoisomerases, Type II; D004875:Ergosterol