阿萨希毛孢子菌响应氟康唑胁迫的蛋白质组分析  

Proteomic analysis of Trichosporon asahii′s response to fluconazole stress

作　　者：杨鑫[1] 夏志宽 敖俊红 祝贺 李季瑾 吴佳敏 徐龄智 杨蓉娅 YANG Xin;XIA Zhikuan;AO Junhong;ZHU He;LI Jijin;WU Jiamin;XU Lingzhi;YANG Rongya(The Seventh Medical Center of PLA General Hospital,Beijing 100010,China)

机构地区：[1]解放军总医院第七医学中心皮肤科,北京100010

出　　处：《中华医院感染学杂志》2025年第6期801-806,共6页Chinese Journal of Nosocomiology

基　　金：国家自然科学基金资助项目(82373494,82073466,81772138);北京市自然科学基金资助项目(7212105,7202201)。

摘　　要：目的探讨氟康唑对阿萨希毛孢子菌(T.asahii)蛋白质组学的影响,从蛋白层面揭示T.asahii对氟康唑胁迫的响应过程及唑类耐药机制。方法以T.asahii AS 2.2174作为研究菌株,基于微量液基稀释法测定氟康唑的最低抑菌浓度(MIC)。应用串联质谱标签(TMT)技术结合液相色谱与串联质谱(LC-MS/MS)技术检测氟康唑(1×MIC)处理前后T.asahii蛋白丰度的变化。以差异倍数≥1.20或≤0.83,且P<0.05作为筛选标准,鉴定差异表达蛋白(DEPs)。对DEPs进行Gene Ontlogy(GO)和京都基因与基因组百科全书(KEGG)富集分析,以了解DEPs的生物学属性以及DEPs参与的主要生物学通路。最后,使用多反应监测(MRM)技术对目标差异蛋白进行靶向验证。结果氟康唑对T.asahii AS 2.2174的MIC值为8μg/ml。共鉴定到DEPs 196个,其中上调DEPs 93个,下调DEPs 103个。功能富集分析提示DEPs主要参与甾醇合成与代谢、药物代谢、应激反应、能量代谢及翻译等生物学过程。目标差异蛋白在MRM靶向验证和TMT-LC-MS/MS检测中具有一致的表达趋势。结论在氟康唑胁迫下,T.asahii蛋白丰度发生显著改变。生物信息学分析揭示了T.asahii响应氟康唑胁迫的复杂分子机制,这些机制对于了解T.asahii唑类耐药性及挖掘新的药物靶点提供了重要见解。OBJECTIVE To explore the effect of fluconazole on proteomics of Trichosporon asahii so as to reveal the responding process of T.asahii to fluconazole stress and the resistance mechanisms to azoles on the protein level.METHODS T.asahii AS 2.2174 was chosen as the research subject,the minimum inhibitory concentration(MIC)of fluconazole was determined by broth microdilution assay.The protein abundance of T.asahii was detected by means of tandem mass tag(TMT)technique combined with liquid chromatography-tandem mass spectrometry(LC-MS/MS)before and after the treatment with fluconazole(1×MIC).The differentially expressed proteins(DEPs)were identified based on the screening standards of fold change≥1.20 or≤0.83 and P<0.05.Gene ontlogy(GO)and Kyoto encyclopedia of genes and genomes(KEGG)enrichment analysis were performed for the DEPs so as to understand the biological property of the DEPs and the major biological pathways that the DEPs involved in.Finally,the targeted validation was carried out for the targeted differentially expressed proteins by using multiple reaction monitoring(MRM).RESULTS The MIC of fluconazole to T.asahii AS 2.2174 was 8μg/ml.Totally 196 DEPs were identified,including 93 upregulated DEPs and 103 downregulated DEPs.The function enrichment analysis showed that the DEPs mainly participated in synthesis and metabolism of sterols,drug metabolism,stress response,energy metabolism and intertranslation.The targeted DEPs showed the consistent expression trends in MRM target validation and TMT-LC-MS/MS.CONCLUSIONS The protein abundance of T.asahii has remarkable change under the fluconazole stress.The bioinformatics analysis reveals the complicated molecular mechanisms of T.asahii in response to the fluconazole stress,which may offer valuable ideas for understanding the drug resistance to azoles and developing new drug targets.

关 键 词：阿萨希毛孢子菌 蛋白质组学 多反应监测 药物胁迫 耐药性 

分 类 号：R379[医药卫生—病原生物学]