Spatial distribution of organelles in leaf cells and soybean root nodules revealed by focused ion beam-scanning electron microscopy
Brandon C. Reagan A , Paul J. -Y. Kim A , Preston D. Perry A , John R. Dunlap B and Tessa M. Burch-Smith A C
+ Author Affiliations
- Author Affiliations
A Department of Biochemistry and Cellular and Molecular Biology, University of Tennessee, Knoxville, 1414 Cumberland Avenue , Knoxville ,TN 37996, USA.
B Advanced Microscopy and Imaging Center, University of Tennessee, Knoxville, 1499 Circle Dr Knoxville, TN 37996, USA.
C Corresponding author. Email: tburchsm@utk.edu
This paper originates from a presentation at the Fourth International Symposium on Plant Signaling and Behavior, Komarov Botanical Institute RAS/Russian Science Foundation, Saint Petersburg, Russia, 19–23 June 2016.
Functional Plant Biology 45(2) 180-191 https://doi.org/10.1071/FP16347
Submitted: 2 October 2016 Accepted: 23 December 2016 Published: 13 February 2017
Abstract
Analysis of cellular ultrastructure has been dominated by transmission electron microscopy (TEM), so images collected by this technique have shaped our current understanding of cellular structure. More recently, three-dimensional (3D) analysis of organelle structures has typically been conducted using TEM tomography. However, TEM tomography application is limited by sample thickness. Focused ion beam-scanning electron microscopy (FIB-SEM) uses a dual beam system to perform serial sectioning and imaging of a sample. Thus FIB-SEM is an excellent alternative to TEM tomography and serial section TEM tomography. Animal tissue samples have been more intensively investigated by this technique than plant tissues. Here, we show that FIB-SEM can be used to study the 3D ultrastructure of plant tissues in samples previously prepared for TEM via commonly used fixation and embedding protocols. Reconstruction of FIB-SEM sections revealed ultra-structural details of the plant tissues examined. We observed that organelles packed tightly together in Nicotiana benthamiana Domin leaf cells may form membrane contacts. 3D models of soybean nodule cells suggest that the bacteroids in infected cells are contained within one large membrane-bound structure and not the many individual symbiosomes that TEM thin-sections suggest. We consider the implications of these organelle arrangements for intercellular signalling.
Additional keywords: FIB-SEM, HPF-FS, membrane continuity, plasmodesmata, root nodules, tomogram.
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