Mehra R K, Thorvaldsen J L, Macreadie I G, Winge D R
Department of Medicine, University of Utah Medical Center, Salt Lake City 84132.
Gene. 1992 May 1;114(1):75-80. doi: 10.1016/0378-1119(92)90709-x.
Candida glabrata harbors multiple genes encoding metallothionein (MT). We have disrupted MT-IIa, an amplified locus, and MT-IIb, a single-copy gene, to determine the roles of various MT genes in CuSO4 resistance in C. glabrata. The concentration of CuSO4 required to inhibit the growth by 50% (IC50) of a C. glabrata strain harboring an amplified MT-IIa locus and a single-copy MT-IIb and MT-I genes was 7 mM in a synthetic complete medium. The IC50 decreased to approx. 1 mM when the amplified MT-IIa locus was deleted. The disruption of the MT-IIb gene decreased the IC50 further to 0.1 mM. The CuSO4 resistance in a strain lacking both of the MT-II genes was attributable to MT-I; no evidence was found for the production of (gamma EC)nG isopeptides. The comparison of the nucleotide sequence of MT-IIb to that of MT-IIa revealed the same coding sequence with differences in the 5' region. However, substantial differences were found in the 3' region. MT-IIb was expressed since we were able to purify the protein from the strain that had an intact MT-IIb gene, but a deleted MT-IIa gene. In addition, CuSO4 resistance was provided by MT-IIb. Northern analysis of the total RNA from varied C. glabrata strains indicated no significant changes in the expression of MT-I in the presence or absence of the MT-II genes.
光滑念珠菌含有多个编码金属硫蛋白（MT）的基因。我们破坏了一个扩增位点MT-IIa和一个单拷贝基因MT-IIb，以确定各种MT基因在光滑念珠菌对硫酸铜抗性中的作用。在合成完全培养基中，对一个含有扩增的MT-IIa位点以及单拷贝MT-IIb和MT-I基因的光滑念珠菌菌株而言，抑制其生长50%（IC50）所需的硫酸铜浓度为7 mM。当扩增的MT-IIa位点被缺失时，IC50降至约1 mM。MT-IIb基因的破坏使IC50进一步降至0.1 mM。在一个同时缺失两个MT-II基因的菌株中，对硫酸铜的抗性归因于MT-I；未发现产生（γEC）nG异肽的证据。MT-IIb与MT-IIa的核苷酸序列比较显示，它们的编码序列相同，但5'区域存在差异。然而，在3'区域发现了显著差异。MT-IIb是有表达的，因为我们能够从一个MT-IIb基因完整但MT-IIa基因缺失的菌株中纯化出该蛋白。此外，MT-IIb赋予了对硫酸铜的抗性。对来自不同光滑念珠菌菌株的总RNA进行的Northern分析表明，无论MT-II基因存在与否，MT-I的表达均无显著变化。
