Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois.
Center for Biophysics and Quantitative Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois.
Traffic. 2019 Nov;20(11):841-850. doi: 10.1111/tra.12683. Epub 2019 Sep 2.
The accumulation of copper in organisms can lead to altered functions of various pathways and become cytotoxic through the generation of reactive oxygen species. In yeast, cytotoxic metals such as Hg , Cd and Cu are transported into the lumen of the vacuole through various pumps. Copper ions are initially transported into the cell by the copper transporter Ctr1 at the plasma membrane and sequestered by chaperones and other factors to prevent cellular damage by free cations. Excess copper ions can subsequently be transported into the vacuole lumen by an unknown mechanism. Transport across membranes requires the reduction of Cu to Cu . Labile copper ions can interact with membranes to alter fluidity, lateral phase separation and fusion. Here we found that CuCl potently inhibited vacuole fusion by blocking SNARE pairing. This was accompanied by the inhibition of V-ATPase H pumping. Deletion of the vacuolar reductase Fre6 had no effect on the inhibition of fusion by copper. This suggests that Cu is responsible for the inhibition of vacuole fusion and V-ATPase function. This notion is supported by the differential effects of chelators. The Cu -specific chelator triethylenetetramine rescued fusion, whereas the Cu -specific chelator bathocuproine disulfonate had no effect on the inhibited fusion.
铜在生物体中的积累会导致各种途径的功能改变，并通过活性氧的产生而产生细胞毒性。在酵母中，汞、镉和铜等细胞毒性金属通过各种泵被运送到液泡腔室中。铜离子最初通过质膜上的铜转运蛋白 Ctr1 被运入细胞，并被伴侣蛋白和其他因素隔离，以防止游离阳离子造成细胞损伤。随后，多余的铜离子可以通过未知的机制被运入液泡腔室。跨膜运输需要将 Cu 还原为 Cu 。不稳定的铜离子可以与膜相互作用，改变流动性、横向相分离和融合。在这里，我们发现 CuCl 通过阻断 SNARE 配对，强烈抑制液泡融合。这伴随着 V-ATPase H 泵的抑制。液泡还原酶 Fre6 的缺失对铜抑制融合没有影响。这表明铜是抑制液泡融合和 V-ATPase 功能的原因。螯合剂的差异效应支持了这一观点。铜特异性螯合剂三乙烯四胺挽救了融合，而铜特异性螯合剂二硫代二氨基二苯并噻唑对受抑制的融合没有影响。
