Institut für Molekulare Infektionsbiologie, Universität Würzburg, Röntgenring 11, D-97070 Würzburg, Germany.
Antimicrob Agents Chemother. 2010 Jan;54(1):353-9. doi: 10.1128/AAC.01102-09. Epub 2009 Nov 2.
The zinc cluster transcription factor Upc2p mediates upregulation of ergosterol biosynthesis genes in response to ergosterol depletion in the fungal pathogen Candida albicans. One mechanism of acquired resistance to the antifungal drug fluconazole, which inhibits ergosterol biosynthesis, is constitutively increased expression of the ERG11 gene encoding the drug target enzyme. A G648D mutation in Upc2p has recently been shown to cause hyperactivity of the transcription factor, resulting in overexpression of ergosterol biosynthesis genes and increased fluconazole resistance. In order to investigate if gain-of-function mutations in Upc2p are a common mechanism of ERG11 upregulation and fluconazole resistance, we sequenced the UPC2 alleles of four ERG11-overexpressing, fluconazole-resistant C. albicans isolates and matched susceptible isolates from the same patients. In three of the isolate pairs, no differences in the UPC2 alleles were found, suggesting that mechanisms other than Upc2p mutations can cause ERG11 overexpression. One resistant isolate had become homozygous for a UPC2 allele containing a G1927A substitution that caused an alanine-to-threonine exchange at amino acid position 643 of Upc2p. Replacement of one of the endogenous UPC2 alleles in a fluconazole-susceptible strain by the UPC2(A643T) allele resulted in ERG11 overexpression and increased fluconazole resistance, which was further elevated when the A643T mutation was also introduced into the second UPC2 allele. These results further establish gain-of-function mutations in UPC2, which can be followed by loss of heterozygosity for the mutated allele, as a mechanism of ERG11 overexpression and increased fluconazole resistance in C. albicans, but other mechanisms of ERG11 upregulation also exist.
锌簇转录因子 Upc2p 介导真菌病原体白色念珠菌中固醇生物合成基因的上调，以响应固醇的耗竭。对抑制固醇生物合成的抗真菌药物氟康唑获得性耐药的一种机制是药物靶酶编码基因 ERG11 的组成型表达增加。最近表明，Upc2p 中的 G648D 突变导致转录因子过度活跃，导致固醇生物合成基因过度表达和氟康唑耐药性增加。为了研究 Upc2p 的功能获得性突变是否是 ERG11 上调和氟康唑耐药的常见机制，我们对四个 ERG11 过表达、氟康唑耐药的白色念珠菌分离株的 UPC2 等位基因进行了测序，并与来自同一患者的敏感分离株进行了匹配。在三对分离株中，未发现 UPC2 等位基因的差异，这表明除了 Upc2p 突变之外，还有其他机制导致 ERG11 过表达。一个耐药分离株的 UPC2 等位基因变成纯合子，其中包含 G1927A 取代，导致 Upc2p 中第 643 位氨基酸的丙氨酸到苏氨酸交换。在氟康唑敏感株中，用 UPC2(A643T)等位基因替换一个内源性 UPC2 等位基因导致 ERG11 过表达和氟康唑耐药性增加，当第二个 UPC2 等位基因也引入 A643T 突变时，耐药性进一步提高。这些结果进一步证实了 UPC2 的功能获得性突变，随后是突变等位基因的杂合性丧失，是白色念珠菌中 ERG11 过表达和氟康唑耐药性增加的一种机制，但也存在其他 ERG11 上调的机制。
