Centre for Infectious Diseases and Microbiology, Westmead Millennium Institute, NSW 2145, Australia.
Fungal Genet Biol. 2013 Dec;61:23-32. doi: 10.1016/j.fgb.2013.09.005. Epub 2013 Sep 26.
Miltefosine (MI) has in vitro fungicidal activity against pathogenic fungi. However, mechanisms of resistance to MI have not been studied. By screening a genomic library of the model yeast, Saccharomyces cerevisiae, we identified HXT13 as a candidate genetic determinant of MI resistance. HXT13 belongs to the yeast hexose transporter family, which mediates hexose sugar uptake and is included in the major facilitator superfamily (MFS). We now report that overexpression of HXT13, but not of the closely-related genes, HXT15 and HXT17, and the more distantly related HXT14, resulted in a stable MI-resistant phenotype in S. cerevisiae. Resistance of the HXT13 overexpressing strain to MI correlated with higher cell viability following MI exposure as assessed by SYTOX® green staining compared with the control and overexpressing HXT14 strains. The mechanism of resistance in the HXT13 overexpressing strain was due to increased ATP-independent MI efflux. However, resistance to MI of the HXT13-overexpressing strain did not extend to other drugs including the echinocandins, amphotericin B, azoles, cycloheximide and sulfometuron methyl, ruling out the involvement of HXT13 in multidrug resistance. In summary, we have identified a new function of the hexose sugar transporter gene HXT13 when overexpressed in S. cerevisiae, namely, in efflux of MI and in mediating MI resistance.
米替福新（MI）在体外对致病性真菌具有杀真菌活性。然而，尚未研究对 MI 产生耐药性的机制。通过筛选模式酵母酿酒酵母的基因组文库，我们鉴定出 HXT13 是 MI 耐药性的候选遗传决定因素。HXT13 属于酵母己糖转运蛋白家族，该家族介导己糖摄取，包含在主要易化因子超家族（MFS）中。我们现在报告，在酿酒酵母中过表达 HXT13，但不是过表达紧密相关的基因 HXT15 和 HXT17，以及更远相关的 HXT14，导致稳定的 MI 耐药表型。与对照和过表达 HXT14 菌株相比，过表达 HXT13 的菌株在暴露于 MI 后通过 SYTOX®绿色染色评估的细胞活力更高，这与 MI 耐药性相关。过表达 HXT13 菌株的耐药机制是由于增加了无 ATP 依赖性 MI 外排。然而，过表达 HXT13 的菌株对 MI 的耐药性并不扩展到其他药物，包括棘白菌素、两性霉素 B、唑类、环己酰亚胺和磺酰脲甲酯，排除了 HXT13 参与多药耐药性的可能性。总之，我们已经确定了在酿酒酵母中过表达的己糖转运蛋白基因 HXT13 的新功能，即 MI 的外排和介导 MI 耐药性。
