转录因子Pho4参与白念珠菌药物敏感性的调控

Transcription factor Pho4 is involved in the regulation of drug susceptibility of Candida albicans

目的通过对缺失相应转录因子基因的白念珠菌进行抗真菌药物敏感性的筛选,考察转录因子对白念珠菌耐药性的影响及调控机制。方法通过微量液基稀释法、点板实验(Spot Assay)检测实验菌株对抗真菌药物的敏感性。采用实时定量PCR(RT-PCR)的方法检测白念珠菌耐药性相关MDR1,CDR1以及ERG11的表达,并通过检测菌株对罗丹明6G的外排能力进一步检测菌株对抗真菌药物的外排能力。结果最低抑菌浓度(minimal inhibitory concentration,MIC)测定和Spot Assay实验结果表明,与亲本菌相比,PHO4基因缺失菌对氟康唑、咪康唑的敏感性显著升高。虽然耐药相关基因的表达增加,但对罗丹明6G的外排能力降低,抗氧化应激能力下降。结论转录因子Pho4的缺失可能通过降低白念珠菌的抗氧化应激能力,减弱对药物的外排作用而导致对唑类药物敏感,但其具体的调控机制有待进一步研究。

Objective To study the influence of Candida albicans transcription factors（TFs）on drug susceptibility and the relevant regulating mechanisms,a screening of a TF deletion mutant library had been conducted.Methods The broth microdilution method and spot assay were used to determine the susceptibility of TF deletion strains.The expression of multidrug resistance genes MDR1,CDR1 and ERG11 were examined by real-time PCR,and the analysis of rhodamine 6Gefflux was performed to evaluate the efflux pump activity.Results The measurement of drug minimal inhibitory concentration and spot assay indicated an increase of susceptibility to fluconazole and miconazole in PHO4 deleted mutant compared to the parental strain.Although the mutant had a higher expression of the drug resistance genes,the amount of rhodamine 6Gefflux and the oxidative stress resistance of it were lower than the parental strain.Conclusion Deletion of PHO4 in C.albicans might cause a decrease in oxidative stress resistance and drug efflux ability,finally result in hypersusceptibility to antifungal drugs.