Boston Biomedical Research Institute, 64 Grove Street, Watertown, MA 02472, USA.
Methods. 2011 Mar;53(3):232-7. doi: 10.1016/j.ymeth.2010.12.001. Epub 2010 Dec 6.
Experiments in yeast have significantly contributed to our understanding of general aspects of biochemistry, genetics, and cell biology. Yeast models have also delivered deep insights in to the molecular mechanism underpinning human diseases, including neurodegenerative diseases. Many neurodegenerative diseases are associated with the conversion of a protein from a normal and benign conformation into a disease-associated and toxic conformation - a process called protein misfolding. The misfolding of proteins with abnormally expanded polyglutamine (polyQ) regions causes several neurodegenerative diseases, such as Huntington's disease and the Spinocerebellar Ataxias. Yeast cells expressing polyQ expansion proteins recapitulate polyQ length-dependent aggregation and toxicity, which are hallmarks of all polyQ-expansion diseases. The identification of modifiers of polyQ toxicity in yeast revealed molecular mechanisms and cellular pathways that contribute to polyQ toxicity. Notably, several of these findings in yeast were reproduced in other model organisms and in human patients, indicating the validity of the yeast polyQ model. Here, we describe different expression systems for polyQ-expansion proteins in yeast and we outline experimental protocols to reliably and quantitatively monitor polyQ toxicity in yeast.
酵母实验极大地促进了我们对生物化学、遗传学和细胞生物学一般方面的理解。酵母模型还深入了解了人类疾病的分子机制，包括神经退行性疾病。许多神经退行性疾病与蛋白质从正常和良性构象转化为与疾病相关的毒性构象有关，这一过程称为蛋白质错误折叠。具有异常扩展的多聚谷氨酰胺（polyQ）区域的蛋白质错误折叠会导致几种神经退行性疾病，如亨廷顿病和脊髓小脑共济失调。表达 polyQ 扩展蛋白的酵母细胞重现了 polyQ 长度依赖性聚集和毒性，这是所有 polyQ 扩展疾病的标志。在酵母中鉴定 polyQ 毒性的修饰因子揭示了导致 polyQ 毒性的分子机制和细胞途径。值得注意的是，酵母中的一些发现也在其他模式生物和人类患者中得到了重现，表明了酵母 polyQ 模型的有效性。在这里，我们描述了酵母中 polyQ 扩展蛋白的不同表达系统，并概述了实验方案，以可靠和定量地监测酵母中的 polyQ 毒性。
