Clinical Mycology Section, Laboratory of Clinical Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Antimicrob Agents Chemother. 2010 Aug;54(8):3308-17. doi: 10.1128/AAC.00535-10. Epub 2010 Jun 14.
DNA microarrays were used to analyze Candida glabrata oropharyngeal isolates from seven hematopoietic stem cell transplant recipients whose isolates developed azole resistance while the recipients received fluconazole prophylaxis. Transcriptional profiling of the paired isolates revealed 19 genes upregulated in the majority of resistant isolates compared to their paired susceptible isolates. All seven resistant isolates had greater than 2-fold upregulation of C. glabrata PDR1 (CgPDR1), a master transcriptional regulator of the pleiotropic drug resistance (PDR) network, and all seven resistant isolates showed upregulation of known CgPDR1 target genes. The altered transcriptome can be explained in part by the observation that all seven resistant isolates had acquired a single nonsynonymous mutation in their CgPDR1 open reading frame. Four mutations occurred in the regulatory domain (L280P, L344S, G348A, and S391L) and one in the activation domain (G943S), while two mutations (N764I and R772I) occurred in an undefined region. Association of azole resistance and the CgPDR1 mutations was investigated in the same genetic background by introducing the CgPDR1 sequences from one sensitive isolate and five resistant isolates into a laboratory azole-hypersusceptible strain (Cgpdr1 strain) via integrative transformation. The Cgpdr1 strain was restored to wild-type fluconazole susceptibility when transformed with CgPDR1 from the susceptible isolate but became resistant when transformed with CgPDR1 from the resistant isolates. However, despite the identical genetic backgrounds, upregulation of CgPDR1 and CgPDR1 target genes varied between the five transformants, independent of the domain locations in which the mutations occurred. In summary, gain-of-function mutations in CgPDR1 contributed to the clinical azole resistance, but different mutations had various degrees of impact on the CgPDR1 target genes.
DNA 微阵列用于分析 7 例接受造血干细胞移植的患者的口腔假丝酵母菌属分离株，这些患者在接受氟康唑预防治疗时其分离株出现唑类药物耐药。与配对的敏感分离株相比，大多数耐药分离株中 19 个基因的转录谱上调。7 个耐药分离株的 C. glabrata PDR1（CgPDR1）均有超过 2 倍的上调，CgPDR1 是多药耐药（PDR）网络的主要转录调节因子，所有 7 个耐药分离株均显示已知的 CgPDR1 靶基因上调。这种改变的转录组可以部分解释为观察到所有 7 个耐药分离株在其 CgPDR1 开放阅读框中均获得了单个非同义突变。4 个突变发生在调节域（L280P、L344S、G348A 和 S391L），1 个突变发生在激活域（G943S），而另外 2 个突变（N764I 和 R772I）发生在未定义区域。通过整合转化，将一个敏感分离株和 5 个耐药分离株的 CgPDR1 序列导入实验室唑类药物低敏菌株（Cgpdr1 菌株），在相同的遗传背景下研究了唑类药物耐药与 CgPDR1 突变的相关性。当用敏感分离株的 CgPDR1 转化 Cgpdr1 菌株时，该菌株恢复为野生型氟康唑敏感性，但当用耐药分离株的 CgPDR1 转化时则变为耐药。然而，尽管遗传背景相同，5 个转化株之间 CgPDR1 和 CgPDR1 靶基因的上调程度不同，与突变发生的结构域位置无关。总之，CgPDR1 的功能获得性突变导致了临床唑类药物耐药，但不同的突变对 CgPDR1 靶基因的影响程度不同。
