Three-dimensional intracellular structure of a whole rice mesophyll cell observed with FIB-SEM
Three-dimensional intracellular structure of a whole rice mesophyll cell observed with FIB-SEM
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DOI：
10.1093/aob/mcx036
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发表时间：
2017-07-01
期刊：
影响因子：
4.2
通讯作者：
Taniguchi, Mitsutaka
中科院分区：
文献类型：
--
作者：
Oi, Takao;Enomoto, Sakiko;Taniguchi, Mitsutaka
利用透射电镜（TEM）对水稻叶片超薄切片进行了二维观察，结果表明，水稻叶片的薄壁组织由叶肉细胞组成，细胞边界复杂，细胞内排列着叶绿体。水稻叶片的分叶细胞形状和叶绿体的位置被认为增加了光合作用气体交换表面的面积。然而，揭示水稻叶肉细胞的三维（3-D）超微结构的细胞图像还没有可视化。在这项研究中，一个完整的水稻叶肉细胞观察使用聚焦离子束扫描电子显微镜（FIB-SEM），它提供了许多连续切片自动，快速和正确的，从而使三维细胞结构reconstruct.Methods水稻叶片化学固定使用常规TEM观察的方法，包埋在树脂中，随后设置在FIB-SEM室。使用FIB对样本块进行横向切片，并使用SEM捕获块面图像。对200 - 500个切片（每个50 nm厚）重复切片和成像过夜。得到的大体积图像堆栈（x = 25 μ m，y = 25 μ m，z = 10-25 μ m）包含一个或两个完整的叶肉细胞。利用图像处理软件重建整个叶肉细胞的三维模型。KeyResults重建的细胞模型呈圆盘状，细胞周围有几个裂片。胞室形状增加了表面积，并且表面积与体积的比率是具有相同体积的圆柱体的两倍。叶绿体占细胞体积的一半，呈片状沿着细胞叶分布，覆盖了大部分细胞内表面，相邻叶绿体紧密相连。结论利用纤维束扫描电镜三维观察了水稻叶片叶肉细胞的细胞和亚细胞超微结构。三维模型和数值信息支持水稻叶肉细胞增加细胞表面和片状叶绿体增强其CO2吸收的假设。
Background and Aims Ultrathin sections of rice leaf blades observed two-dimensionally using a transmission electron microscope (TEM) show that the chlorenchyma is composed of lobed mesophyll cells, with intricate cell boundaries, and lined with chloroplasts. The lobed cell shape and chloroplast positioning are believed to enhance the area available for the gas exchange surface for photosynthesis in rice leaves. However, a cell image revealing the three-dimensional (3-D) ultrastructure of rice mesophyll cells has not been visualized. In this study, a whole rice mesophyll cell was observed using a focused ion beam scanning electron microscope (FIB-SEM), which provides many serial sections automatically, rapidly and correctly, thereby enabling 3-D cell structure reconstruction.Methods Rice leaf blades were fixed chemically using the method for conventional TEM observation, embedded in resin and subsequently set in the FIB-SEM chamber. Specimen blocks were sectioned transversely using the FIB, and block-face images were captured using the SEM. The sectioning and imaging were repeated overnight for 200500 slices (each 50nm thick). The resultant large-volume image stacks (x = 25 mu m, y = 25 mu m, z = 10-25 mu m) contained one or two whole mesophyll cells. The 3-D models of whole mesophyll cells were reconstructed using image processing software.Key Results The reconstructed cell models were discoid shaped with several lobes around the cell periphery. The cell shape increased the surface area, and the ratio of surface area to volume was twice that of a cylinder having the same volume. The chloroplasts occupied half the cell volume and spread as sheets along the cell lobes, covering most of the inner cell surface, with adjacent chloroplasts in close contact with each other.Conclusions Cellular and sub-cellular ultrastructures of a whole mesophyll cell in a rice leaf blade are demonstrated three-dimensionally using a FIB-SEM. The 3-D models and numerical information support the hypothesis that rice mesophyll cells enhance their CO2 absorption with increased cell surface and sheet-shaped chloroplasts.