Department of Microbiology and Virology, Faculty of Natural Sciences, Comenius University in Bratislava, Slovak Republic.
Yeast. 2010 May;27(5):277-84. doi: 10.1002/yea.1751.
The PDR3 gene encodes one of the main transcriptional activators involved in the control of multidrug resistance in the yeast Saccharomyces cerevisiae. Recently, it has been demonstrated that a specific D853Y mutation results in the loss of transactivation activity of Pdr3p and its conversion to multicopy suppressor of multidrug resistance. In this study, the Asp853 in Pdr3p was replaced by eight different amino acids and the function of mutated proteins was analysed. Different levels of complementation of cycloheximide hypersensitivity and expression of autoregulated PDR3 and its PDR5 target in the pdr1Deltapdr3Delta mutant strain, ranging from that of the wild-type to loss-of-function alleles, were observed in pdr3 mutants containing Pro, Glu, Arg, Asn, Ser, Leu, Phe, Ile or Tyr instead of Asp853 in Pdr3p. The introduction of the D853Y mutation into gain-of-function Pdr3p suppressed the transcription of the PDR3 and PDR5 genes and reduced both the rhodamine 6G efflux rate and the drug resistance level in corresponding double mutants. The results indicate that, while Pdr3p can tolerate several substitutions of Asp853, the occurrence of a hydrophobic amino acid at this position has an adverse effect on its function.
PDR3 基因编码酵母酿酒酵母中多药耐药性控制的主要转录激活因子之一。最近，已经证明特定的 D853Y 突变导致 Pdr3p 的转录激活活性丧失，并将其转化为多拷贝耐药性的多拷贝抑制物。在这项研究中，将 Pdr3p 中的天冬氨酸 853 替换为八种不同的氨基酸，并分析了突变蛋白的功能。在包含 Pro、Glu、Arg、Asn、Ser、Leu、Phe、Ile 或 Tyr 而不是 Pdr3p 中的 Asp853 的 pdr3 突变体中，观察到不同水平的环丝氨酸超敏性互补和自身调节的 PDR3 和其 PDR5 靶基因在 pdr1Deltapdr3Delta 突变体中的表达，从野生型到功能丧失等位基因不等。将 D853Y 突变引入功能获得性 Pdr3p 中，抑制了 PDR3 和 PDR5 基因的转录，并降低了相应双突变体中的罗丹明 6G 外排率和药物抗性水平。结果表明，虽然 Pdr3p 可以容忍 Asp853 的几种取代，但该位置上出现疏水性氨基酸对其功能有不利影响。
