Laboratory of Plant Production Chemistry, Department of Applied Biology and Chemistry, Tokyo University of Agriculture, Tokyo 156-8502, Japan.
Plant Cell Environ. 2010 Feb;33(2):174-87. doi: 10.1111/j.1365-3040.2009.02068.x. Epub 2009 Nov 11.
To evaluate Ni dynamics at the subcellular level, the distribution and speciation of Ni were determined in wild-type (WT) and Ni-tolerant (NIT) tobacco BY-2 cell lines. When exposed to low but toxic levels of Ni, NIT cells were found to contain 2.5-fold more Ni (14% of whole-cell Ni values) in their cell walls than WT cells (6% of whole-cell Ni values). In addition to higher levels of Ni in the apoplast, a higher proportion (94%) of symplastic Ni was localized in the vacuoles of NIT cells than in the vacuoles of WT cells (81%). The concentration of cytosolic Ni in the NIT cells was significantly lower (18 nmol g(-1) FW) than that in the WT cells (85 nmol g(-1) FW). In silico simulation showed that 95% of vacuolar Ni was in the form of Ni-citrate complexes, and that free Ni(2+) was virtually absent in the NIT cells. On the other hand, the amount of free metal ions was markedly increased in WT cells because free citrate was depleted by chelation of Ni. A protoplast viability assay using BCECF-AM further demonstrated that the main mechanism that confers strong Ni tolerance was present in the symplast as opposed to the cell wall.
为了评估亚细胞水平的镍动态，在野生型（WT）和镍耐受（NIT）烟草 BY-2 细胞系中确定了镍的分布和形态。当暴露于低但有毒水平的镍时，与 WT 细胞（全细胞镍值的 6%）相比，NIT 细胞的细胞壁中镍含量多 2.5 倍（全细胞镍值的 14%）。除了细胞外间隙中镍含量较高外，与 WT 细胞（81%）相比，NIT 细胞中质体镍的比例（94%）更高地定位于液泡中。NIT 细胞中胞质镍的浓度明显较低（18 nmol g(-1) FW），而 WT 细胞中胞质镍的浓度较高（85 nmol g(-1) FW）。计算机模拟表明，95%的液泡镍以柠檬酸镍复合物的形式存在，而 NIT 细胞中几乎不存在游离的 Ni(2+)。另一方面，由于镍的螯合耗尽了游离柠檬酸，WT 细胞中游离金属离子的数量明显增加。使用 BCECF-AM 的原生质体活力测定进一步表明，赋予强镍耐受性的主要机制存在于质体中，而不是细胞壁中。
