Department of Biological Science, Graduate School of Science, Hiroshima University, Higashi-Hiroshima, Hiroshima, 739-8526, Japan.
Genes Cells. 2012 Jul;17(7):597-610. doi: 10.1111/j.1365-2443.2012.01612.x. Epub 2012 Jun 12.
Agrobacterium strains containing a Ti plasmid can transfer T-DNA not only to plants but also to fungi, including the yeast Saccharomyces cerevisiae. However, no Agrobacterium strain harboring an Ri plasmid has been evaluated in fungal transformation. Some Ri plasmids have GALLS , instead of virE1 and virE2. GALLS protein can functionally substitute in plant transformation for a structurally different protein VirE2. In this study, we compared the yeast transformation ability among Agrobacterium donors: a strain containing a Ti plasmid, strains harboring either an agropine-type or a mikimopine-type Ri plasmid, and a strain having a modified Ri plasmid supplemented with a Ti plasmid type virE operon. Agrobacterium strains possessing GALLS transformed yeast cells far less efficiently than the strain containing virE operon. Production of GALLS in recipient yeast cells improved the yeast transformation mediated by an Agrobacterium strain lacking neither GALLS nor virE operon. A reporter assay to detect mobilization of the proteins fused with Cre recombinase revealed that VirE2 protein is much more abundant in yeast cells than GALLS. Based on these results, we concluded that the low yeast transformability mediated by Agrobacterium strains having the Ri plasmid is because of low amount of mobilized GALLS in yeast cells.
含有 Ti 质粒的根瘤农杆菌不仅可以将 T-DNA 转移到植物上，还可以转移到真菌上，包括酵母酿酒酵母。然而，尚未评估含有 Ri 质粒的根瘤农杆菌菌株在真菌转化中的作用。一些 Ri 质粒具有 GALLS，而不是 virE1 和 virE2。GALLS 蛋白可以在植物转化中替代结构不同的蛋白 VirE2。在这项研究中，我们比较了含有 Ti 质粒的根瘤农杆菌菌株、携带农杆菌型或米克莫皮农型 Ri 质粒的菌株以及携带改良 Ri 质粒并补充 Ti 质粒型 virE 操纵子的菌株之间的酵母转化能力。含有 GALLS 的根瘤农杆菌菌株转化酵母细胞的效率远低于含有 virE 操纵子的菌株。在既不缺乏 GALLS 也不缺乏 virE 操纵子的根瘤农杆菌菌株介导的酵母转化中，GALLS 在受者酵母细胞中的产生提高了转化效率。检测与 Cre 重组酶融合的蛋白质的转移动力学的报告测定表明，VirE2 蛋白在酵母细胞中的丰度远高于 GALLS。基于这些结果，我们得出结论，含有 Ri 质粒的根瘤农杆菌菌株介导的低酵母转化能力是由于酵母细胞中可动员的 GALLS 数量较少所致。
