Abstract
Due to their position firmly anchored within the plant cell wall, plasmodesmata (PD) are notoriously difficult to isolate from plant tissue. Yet, getting access to isolated PD represents the most straightforward strategy for the identification of their molecular components. Proteomic and lipidomic analyses of such PD fractions have provided and will continue to provide critical information on the functional and structural elements that define these membranous nano-pores. Here, we describe a two-step simple purification procedure that allows isolation of pure PD-derived membranes from Arabidopsis suspension cells. The first step of this procedure consists in isolating cell wall fragments containing intact PD while free of contamination from other cellular compartments. The second step relies on an enzymatic degradation of the wall matrix and the subsequent release of “free” PD. Isolated PD membranes provide a suitable starting material for the analysis of PD-associated proteins and lipids.
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References
Salmon MS, Bayer EM (2013) Dissecting plasmodesmata molecular composition by mass spectrometry-based proteomics. Front Plant Sci 3:307
Faulkner C, Maule A (2011) Opportunities and successes in the search for plasmodesmal proteins. Protoplasma 248:27–38
Sagi G, Katz A, Guenoune-Gelbart D et al (2005) Class 1 reversibly glycosylated polypeptides are plasmodesmal-associated proteins delivered to plasmodesmata via the golgi apparatus. Plant Cell 17:1788–1800
Lee JY, Taoka K, Yoo BC et al (2005) Plasmodesmal-associated protein kinase in tobacco and Arabidopsis recognizes a subset of non-cell-autonomous proteins. Plant Cell 17: 2817–2831
Lee JY, Yoo BC, Rojas MR et al (2003) Selective trafficking of non-cell-autonomous proteins mediated by NtNCAPP1. Science 299:392–396
Simpson C, Thomas C, Findlay K et al (2009) An Arabidopsis GPI-anchor plasmodesmal neck protein with callose binding activity and potential to regulate cell-to-cell trafficking. Plant Cell 21:581–594
Thomas CL, Bayer EM, Ritzenthaler C et al (2008) Specific targeting of a plasmodesmal protein affecting cell-to-cell communication. PLoS Biol 6:e7
Jo Y, Cho WK, Rim Y et al (2011) Plasmodesmal receptor-like kinases identified through analysis of rice cell wall extracted proteins. Protoplasma 248:191–203
Levy A, Erlanger M, Rosenthal M et al (2007) A plasmodesmata-associated beta-1,3-glucanase in Arabidopsis. Plant J 49:669–682
Epel BL, Kuchuck B, Kotlizky G et al (1995) Isolation and characterization of plasmodesmata. Methods Cell Biol 50:237–253
Faulkner C, Petutschnig E, Benitez-Alfonso Y et al (2013) LYM2-dependent chitin perception limits molecular flux via plasmodesmata. Proc Natl Acad Sci U S A 110: 9166–9170
Fernandez-Calvino L, Faulkner C, Walshaw J et al (2011) Arabidopsis plasmodesmal proteome. PLoS One 6:e18880
Benitez-Alfonso Y, Faulkner C, Pendle A et al (2013) Symplastic intercellular connectivity regulates lateral root patterning. Dev Cell 26: 136–147
Bayer E, Thomas CL, Maule AJ (2004) Plasmodesmata in Arabidopsis thaliana suspension cells. Protoplasma 223:93–102
Hofte H, Chrispeels MJ (1992) Protein sorting to the vacuolar membrane. Plant Cell 4: 995–1004
Vallon O, Wollman FA, Olive J (1986) Lateral distribution of the main protein complexes of the photosynthetic apparatus in Chlamydomonas reinhardtii and in spinach: an immunocytochemical study using intact thylakoid membranes and PS II enriched membrane preparation. J Photobiochem Photobiophys 12:203–220
Morsomme P, Dambly S, Maudoux O et al (1998) Single point mutations distributed in 10 soluble and membrane regions of the Nicotiana plumbaginifolia plasma membrane PMA2 H + -ATPase activate the enzyme and modify the structure of the C-terminal region. J Biol Chem 273:34837–34842
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Grison, M.S., Fernandez-Calvino, L., Mongrand, S., Bayer, E.M.F. (2015). Isolation of Plasmodesmata from Arabidopsis Suspension Culture Cells. In: Heinlein, M. (eds) Plasmodesmata. Methods in Molecular Biology, vol 1217. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1523-1_5
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DOI: https://doi.org/10.1007/978-1-4939-1523-1_5
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