Abstract
In plants as well as other organisms, protein localization alone is insufficient to provide a mechanistic link between stimulus and process regulation. This is because protein–protein interactions are central to the regulation of biological processes. However, they remain very difficult to detect in situ, with the choice of tools for the detection of protein–protein interaction in situ still in need of expansion. Here, we provide a protocol for the detection and accurate localization of protein interactions based on the combination of a whole-mount proximity ligation assay and iRoCS, a coordinate system able to standardize subtle differences between the architecture of individual Arabidopsis roots.
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References
Uhlen M, Ponten F (2005) Antibody-based proteomics for human tissue profiling. Mol Cell Proteomics 4:384–393
Sprinzak E, Sattath S, Margalit H (2003) How reliable are experimental protein–protein interaction data? J Mol Biol 327:919–923
Hu CD, Chinenov Y, Kerppola TK (2002) Visualization of interactions among bZip and Rel family proteins in living cells using bimolecular fluorescence complementation. Mol Cell 9:789–798
Bartel PL, Fields S (1995) Analyzing protein–protein initeractions using yeast 2-hybrid system. Oncogene Tech 254:241–263
Soderberg O, Gullberg M, Jarvius M, Ridderstrale K, Leuchowius KJ, Jarvius J et al (2006) Direct observation of individual endogenous protein complexes in situ by proximity ligation. Nat Methods 3:995–1000
Soderberg O, Leuchowius KJ, Gullberg M, Jarvius M, Weibrecht I, Larsson LG et al (2008) Characterizing proteins and their interactions in cells and tissues using the in situ proximity ligation assay. Methods 45:227–232
Apelt F, Breuer D, Nikoloski Z, Stitt M, Kragler F (2015) Phytotyping(4D): a light-field imaging system for non-invasive and accurate monitoring of spatio-temporal plant growth. Plant J 82:693–706
Fernandez R, Das P, Mirabet V, Moscardi E, Traas J, Verdeil JL et al (2010) Imaging plant growth in 4D: robust tissue reconstruction and lineaging at cell resolution. Nat Methods 7:547–U594
Montenegro-Johnson TD, Stamm P, Strauss S, Topham AT, Tsagris M, Wood ATA et al (2015) Digital single-cell analysis of plant organ development using 3DCellAtlas. Plant Cell 27:1018–1033
de Reuille PB, Routier-Kierzkowska AL, Kierzkowski D, Bassel GW, Schupbach T, Tauriello G et al (2015) MorphoGraphX: a platform for quantifying morphogenesis in 4D. Elife 4:05864
Schmidt T, Pasternak T, Liu K, Blein T, Aubry-Hivet D, Dovzhenko A et al (2014) The iRoCS toolbox: 3D analysis of the plant root apical meristem at cellular resolution. Plant J 77:806–814
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Pasternak, T., Teale, W., Falk, T., Ruperti, B., Palme, K. (2018). A PLA-iRoCS Pipeline for the Localization of Protein–Protein Interactions In Situ. In: Wagner, B. (eds) Phenotypic Screening. Methods in Molecular Biology, vol 1787. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7847-2_12
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DOI: https://doi.org/10.1007/978-1-4939-7847-2_12
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-7846-5
Online ISBN: 978-1-4939-7847-2
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