Holland Sara L, Avery Simon V
School of Biology, Institute of Genetics, The University of Nottingham, University Park, Nottingham, UK.
Toxicol Sci. 2009 Oct;111(2):437-46. doi: 10.1093/toxsci/kfp170. Epub 2009 Jul 23.
Chromate toxicity is well documented, but the underlying toxic mechanism(s) has yet to be fully elucidated. Following a Cr toxicity screen against > 6000 heterozygous yeast mutants, here we show that Cr resistance requires normal function of the cortical actin cytoskeleton. Furthermore, Cr-stressed yeast cells exhibited an increased number of actin patches, the sites of endocytosis. This was coincident with a marked stimulation of endocytosis following Cr exposure. Genetic dissection of actin nucleation from endocytosis revealed that endocytosis, specifically, was required for Cr resistance. A series of further endocytosis mutants (sac6Delta, chc1Delta, end3Delta) exhibited elevated Cr sensitivity. These mutants also showed markedly elevated cellular Cr accumulation, explaining their sensitivities. In wild-type cells, an initial endocytosis-independent phase of Cr uptake was followed by an endocytosis-dependent decline in Cr accumulation. The results indicate that actin-mediated endocytosis is required to limit Cr accumulation and toxicity. It is proposed that this involves ubiquitin-dependent endocytic inactivation of a plasma membrane Cr transporter(s). We showed that such an action was not dependent on the transporters that have been characterized to date, the sulfate (and chromate) permeases Sul1p and Sul2p.
铬酸盐毒性已有充分记载，但潜在的毒性机制尚未完全阐明。在针对6000多个杂合酵母突变体进行铬毒性筛选后，我们在此表明，对铬的抗性需要皮层肌动蛋白细胞骨架的正常功能。此外，铬胁迫的酵母细胞中肌动蛋白斑点（即内吞作用位点）的数量增加。这与铬暴露后内吞作用的显著刺激同时发生。从内吞作用对肌动蛋白成核进行基因剖析发现，具体而言，内吞作用是铬抗性所必需的。一系列进一步的内吞作用突变体（sac6Delta、chc1Delta、end3Delta）对铬的敏感性升高。这些突变体还显示细胞内铬积累明显增加，这解释了它们的敏感性。在野生型细胞中，铬摄取的初始阶段不依赖内吞作用，随后是依赖内吞作用的铬积累下降。结果表明，肌动蛋白介导的内吞作用对于限制铬积累和毒性是必需的。据推测，这涉及质膜铬转运蛋白的泛素依赖性内吞失活。我们表明，这种作用不依赖于迄今为止已鉴定的转运蛋白，即硫酸盐（和铬酸盐）通透酶Sul1p和Sul2p。
