Studies on mechanism of toxicity of methylmercury

甲基汞毒性机制研究

基本信息

批准号：
15209004
负责人：
NAGANUMA Akira
金额：
$ 32.45万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15209004/
关键词：
methylmercury toxicity resistance ubiquitin F-box proteins Cdc34 Rad23 酵母

项目摘要

We have identified a gene, CDC34 that confers resistance to methylmercury in Saccharomyces cerevisiae by screening a yeast genomic DNA library. CDC34 encodes one of the ubiquitin-conjugating enzymes (Cdc34) that catalyze ubiquitin transfer onto substrate proteins, an important step for protein degradation via ubiquitin-proteasome pathway. In yeast cells, the ubiquitin-conjugating activity of Cdc34 is essential for the Cdc34-mediated resistance to methylmercury. It is possible to conclude that proteins involved in expression of methylmercury toxicity might exist in the cells and the proteins are degraded after its ubiquitination in proteasomes. In ubiquitin system, substrate proteins are recognized by F-box proteins and ubiquitinated by ubiquitin-conjugating enzymes. Thus, we searched F-box proteins which confer resistance to methylmercury, and found that Hrt3 and Ylr224w have role in protection against methylmercury toxicity. Proteins that bind to these F-box proteins might have activity to intensify the toxicity of methylmercury. We identified three proteins that bind to the F-box proteins, Hrt3 and Ylr224w. Overexpression of the genes for these proteins conferred a hyper susceptibility to methylmercury. Moreover, these proteins are found to be ubiquitinated in the cells. These results suggest that the F-box protein-bounding proteins identified by the present study have significant roles in expression of methylmercury toxicity and are decomposed by the ubiquitin-proteasome pathway. Two of these proteins are enzymes involved in synthesis of pyruvate, and overexpression of these proteins might increase cellular concentration of pyruvate. Addition of excess pyruvate into the culture medium significantly prevents toxicity of methylmercury. Increase in cellular production of pyruvate might have an important role in enhancement of toxicity of methylmercury.

我们已经确定了一种基因Cdc34，该基因通过筛选酵母基因组DNA文库来赋予酿酒酵母中甲基汞的耐药性。 CDC34编码一种泛素 - 偶联酶（CDC34），该酶将泛素转移催化至底物蛋白上，这是通过泛素蛋白促蛋白途径蛋白质降解的重要步骤。在酵母细胞中，Cdc34的泛素结合活性对于CDC34介导的对甲基汞的抗性至关重要。可以得出结论，细胞中可能存在参与甲基汞毒性表达的蛋白质，并且蛋白质在蛋白酶体中泛素化后会降解蛋白质。在泛素系统中，底物蛋白被F-box蛋白识别，并通过泛素 - 偶联酶泛素化。因此，我们搜索了赋予甲基汞耐药性的F-box蛋白，发现HRT3和YLR224W在防御甲基汞毒性中起作用。与这些F-box蛋白结合的蛋白质可能具有活性，以加强甲基汞的毒性。我们确定了与F-box蛋白HRT3和YLR224W结合的三种蛋白质。这些蛋白质的基因过表达赋予了对甲基马克里的过度敏感性。此外，发现这些蛋白质在细胞中被泛素化。这些结果表明，本研究鉴定出的含F-box蛋白质的蛋白质在甲基马凝毒性的表达中具有重要作用，并且被泛素 - 蛋白酶体途径分解。这些蛋白中的两种是参与丙酮酸合成的酶，这些蛋白质的过表达可能会增加丙酮酸的细胞浓度。将过量的丙酮酸添加到培养基中可以显着防止甲基质的毒性。丙酮酸细胞产生的增加可能在增强甲基质固醇的毒性方面具有重要作用。