Yokota E, Yukawa C, Muto S, Sonobe S, Shimmen T
Department of Life Science, Faculty of Science, Himeji Institute of Technology, Harima Science Park City, Hyogo 678-12, Japan.
Plant Physiol. 1999 Oct;121(2):525-34. doi: 10.1104/pp.121.2.525.
We have isolated a myosin (referred to as 170-kD myosin) from lily pollen tubes, which consists of 170-kD heavy chain and calmodulin (CaM) light chain and is responsible for cytoplasmic streaming. A 170-kD polypeptide that has similar antigenicity to the 170-kD myosin heavy chain of lily pollen tubes was also present in cultured tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells, and possessed the ability to interact with F-actin in an ATP-dependent manner. In addition to this myosin, we identified biochemically another kind of myosin in BY-2 cells. This myosin consisted of a CaM light chain and a 175-kD heavy chain with antigenicity different from the 170-kD myosin heavy chain. In the present study, we referred to this myosin as 175-kD myosin. This myosin was able to translocate rhodamine-phalloidin (RP)-labeled F-actin at an average velocity of about 9 &mgr;m/s in the motility assay in vitro. In contrast, the sliding velocity of RP-labeled F-actin translocated by fractions containing the 170-kD myosin was 3 to 4 &mgr;m/s. The velocity of cytoplasmic streaming in living BY-2 cells ranged from 2 to 9 &mgr;m/s. The motile activity of 175-kD myosin in vitro was inhibited by Ca(2+) at concentrations higher than 10(-6) M. Immunoblot analyses using an antiserum against the heavy chain of 170- or 175-kD myosin revealed that in tobacco plants, the 175-kD myosin was expressed in leaf, stem, and root, but not in germinating pollen, while 170-kD myosin was present in all of these plant parts and in germinating pollen. These results suggest that the two types of myosins, 170 and 175 kD, presumably participate in cytoplasmic streaming in BY-2 cells and other somatic cells of tobacco plants.
我们从百合花粉管中分离出一种肌球蛋白（称为170-kD肌球蛋白），它由170-kD重链和钙调蛋白（CaM）轻链组成，负责细胞质流动。在培养的烟草（Nicotiana tabacum）亮黄-2（BY-2）细胞中也存在一种与百合花粉管170-kD肌球蛋白重链具有相似抗原性的170-kD多肽，并且它具有以ATP依赖的方式与F-肌动蛋白相互作用的能力。除了这种肌球蛋白外，我们还通过生化方法在BY-2细胞中鉴定出另一种肌球蛋白。这种肌球蛋白由CaM轻链和一条175-kD重链组成，其抗原性与170-kD肌球蛋白重链不同。在本研究中，我们将这种肌球蛋白称为175-kD肌球蛋白。在体外运动分析中，这种肌球蛋白能够以约9μm/s的平均速度转运罗丹明-鬼笔环肽（RP）标记的F-肌动蛋白。相比之下，含有170-kD肌球蛋白的组分转运RP标记的F-肌动蛋白的滑动速度为3至4μm/s。活的BY-2细胞中细胞质流动的速度范围为2至9μm/s。浓度高于10^(-6) M的Ca(2+)会抑制175-kD肌球蛋白在体外的运动活性。使用针对170-或175-kD肌球蛋白重链的抗血清进行免疫印迹分析表明，在烟草植株中，175-kD肌球蛋白在叶、茎和根中表达，但在萌发的花粉中不表达，而170-kD肌球蛋白存在于所有这些植物部位以及萌发的花粉中。这些结果表明，170-kD和175-kD这两种类型的肌球蛋白可能参与了烟草植株BY-2细胞和其他体细胞中的细胞质流动。
