Kawai Shigeyuki, Phan Tuan Anh, Kono Emi, Harada Kazuo, Okai Chihiro, Fukusaki Eiichiro, Murata Kousaku
Department of Basic and Applied Molecular Biotechnology, Division of Food and Biological Science, Graduate School of Agriculture, Kyoto University, Uji, Kyoto, Japan.
J Basic Microbiol. 2009 Feb;49(1):73-81. doi: 10.1002/jobm.200800123.
Intact cells of the yeast Saccharomyces cerevisiae were transformed with exogenous DNA by incubating the cells with plasmid in the presence of polyethylene glycol (PEG), which has been shown to be required, although the underlying has not been elucidated. In this study, we found that incubation of the S. cerevisiae cells with PEG was not only required for the PEG-dependent transformation but also enhanced transformation, suggesting that PEG might cause an intracellular response. To understand the response, microarray and metabolome analyses were conducted. We found that incubation of the cells without PEG caused up-regulation of several genes, including those which are involved in carbon source metabolisms, e.g. fatty acid metabolism, yielding acetyl-CoA and those involved in stress-response. Contrary to this, incubation of the cells with PEG gave no transcriptional change. These microarray data were supported by the results of metabolome analysis for anionic metabolites, implying that the physical effect of PEG on cell membrane, rather than the effect of PEG itself on the intracellular response, could cause high transformation in the PEG-dependent transformation.
将酿酒酵母的完整细胞与质粒在聚乙二醇（PEG）存在的情况下孵育，可实现外源性DNA对细胞的转化。虽然尚未阐明其潜在机制，但PEG已被证明是实现转化所必需的。在本研究中，我们发现酿酒酵母细胞与PEG孵育不仅是PEG依赖性转化所必需的，而且还能增强转化效果，这表明PEG可能引发细胞内反应。为了解该反应，我们进行了微阵列和代谢组分析。我们发现，在无PEG的情况下孵育细胞会导致几个基因上调，包括参与碳源代谢的基因，如产生乙酰辅酶A的脂肪酸代谢相关基因以及参与应激反应的基因。与此相反，在有PEG的情况下孵育细胞则不会引起转录变化。阴离子代谢物的代谢组分析结果支持了这些微阵列数据，这意味着在PEG依赖性转化中，PEG对细胞膜的物理作用而非PEG本身对细胞内反应的作用可能导致高转化率。
