Abstract
Electron tomography is a powerful tool for visualizing subcellular organelles. With the advances in cryo-fixation techniques, it is now possible to reconstruct complex structures in cells preserved close to their native states in three-dimension (3D) using electron tomography. In order to better visualize these objects, 3D models are made from outlines of organelles in individual tomographic slices, which can be used to display morphological features and quantify structural parameters. While outlines of simple organelles can be drawn by hand fairly quickly, it is possible to accelerate 3D modeling of more complex organelles by means of semiautomatic segmentation. In this chapter, we use the example of reconstructing Golgi cisternae of a plant cell into 3D models using the semiautomatic protocol.
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References
Staehelin LA, Kang B-H (2008) Nanoscale architecture of endoplasmic reticulum export sites and of Golgi membranes as determined by electron tomography. Plant Physiol 147:1454–1468. doi:10.1104/pp.108.120618
Donohoe BS, Kang B-H, Gerl MJ et al (2013) Cis-Golgi cisternal assembly and biosynthetic activation occur sequentially in plants and algae. Traffic 14:551–567. doi:10.1111/tra.12052
Karahara I, Kang B-H (2014) High-pressure freezing and low-temperature processing of plant tissue samples for electron microscopy. Methods Mol Biol 1080:147–157. doi:10.1007/978-1-62703-643-6_12
Zhou Q, Li H, Li H et al (2016) Mitochondrial endonuclease G mediates breakdown of paternal mitochondria upon fertilization. Science 353:394–399
Wang Y, Zhang Y, Chen L et al (2016) Kinetics and specificity of paternal mitochondrial elimination in Caenorhabditis elegans. Nat Commun 7:12569. doi:10.1038/ncomms12569
Engel BD, Schaffer M, Albert S et al (2015) In situ structural analysis of Golgi intracisternal protein arrays. Proc Natl Acad Sci U S A 112:11264–11269. doi:10.1073/pnas.1515337112
Engel BD, Schaffer M, Kuhn Cuellar L et al (2015) Native architecture of the Chlamydomonaschloroplast revealed by in situ cryo-electron tomography. Elife 4:3583–3529. doi:10.7554/eLife.04889
Daum B, Kuhlbrandt W (2011) Electron tomography of plant thylakoid membranes. J Exp Bot 62:2393–2402. doi:10.1093/jxb/err034
Kang B-H, Nielsen E, Preuss ML et al (2011) Electron Tomography of RabA4b- and PI-4Kβ1-Labeled Trans Golgi Network Compartments in Arabidopsis. Traffic 12:313–329. doi:10.1111/j.1600-0854.2010.01146.x
Austin JR, Staehelin LA (2011) Three-dimensional architecture of grana and stroma thylakoids of higher plants as determined by electron tomography. Plant Physiol 155:1601–1611. doi:10.1104/pp.110.170647
Toyooka K, Kang B-H (2014) Reconstructing Plant Cells in 3D by Serial Section Electron Tomography. Methods Mol Biol 1080:159–170. doi:10.1007/978-1-62703-643-6_13
Kang B-H (2010) Electron microscopy and high-pressure freezing of Arabidopsis. Methods Cell Biol 96:259–283. doi:10.1016/S0091-679X(10)96012-3
Donohoe B, Mogelsvang S, Staehelin L (2006) Electron tomography of ER, Golgi and related membrane systems. Methods 39:154–162
Kang B-H, Staehelin LA (2008) ER-to-Golgi transport by COPII vesicles in Arabidopsis involves a ribosome-excluding scaffold that is transferred with the vesicles to the Golgi matrix. Protoplasma 234:51–64. doi:10.1007/s00709-008-0015-6
Chklovskii DB, Vitaladevuni S, Scheffer LK (2010) Semi-automated reconstruction of neural circuits using electron microscopy. Curr Opin Neurobiol 20:667–675. doi:10.1016/j.conb.2010.08.002
Jones C, Liu T, Cohan NW et al (2015) Efficient semi-automatic 3D segmentation for neuron tracing in electron microscopy images. J Neurosci Methods 246:13–21. doi:10.1016/j.jneumeth.2015.03.005
Hoang TV, Kizilyaprak C, Spehner D et al (2017) Automatic segmentation of high pressure frozen and freeze-substituted mouse retina nuclei from FIB-SEM tomograms. J Struct Biol 197(2):134–123. doi:10.1016/j.jsb.2016.10.005
Kang B-H (2016) STEM Tomography Imaging of Hypertrophied Golgi Stacks in Mucilage-Secreting Cells. Methods Mol Biol 1496:55–62. doi:10.1007/978-1-4939-6463-5_5
Acknowledgments
This work was supported by grants from the Research Grants Council of Hong Kong (GRF-14126116, AoE/M-05/12, C4011-14R) and the Cooperative Research Program for Agriculture Science and Technology Development (Project No. PJ010953092015) from the Rural Development Administration, Republic of Korea.
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Mai, K.K.K., Kang, BH. (2017). Semiautomatic Segmentation of Plant Golgi Stacks in Electron Tomograms Using 3dmod. In: Jiang, L. (eds) Plant Protein Secretion. Methods in Molecular Biology, vol 1662. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7262-3_8
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DOI: https://doi.org/10.1007/978-1-4939-7262-3_8
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