Multicolor bimolecular fluorescence complementation (mcBiFC) reveals simultaneous formation of alternative CBL/CIPK complexes in planta Export

@article{citeulike:3055858, abstract = {The specificity of intracellular signaling and developmental patterning in biological systems relies on selective interactions between different proteins in specific cellular compartments. The identification of such protein-protein interactions is essential for unraveling complex signaling and regulatory networks. Recently, bimolecular fluorescence complementation (BiFC) has emerged as a powerful technique for the efficient detection of protein interactions in their native subcellular localization. Here we report significant technical advances in plant BiFC methodology. We describe a series of versatile BiFC vector sets that are fully compatible to previously generated vectors (Walter et al., 2004). The new vectors enable the generation of both C-terminal and N-terminal fusion proteins and carry optimized fluorescent protein genes that considerably improve the sensitivity of BiFC. Using these vectors, we describe a multicolor BiFC (mcBiFC) approach for the simultaneous visualization of multiple protein interactions in the same cell. Application to a protein interaction network acting in calcium-mediated signal transduction revealed the concurrent interaction of the protein kinase CIPK24 with the calcium sensors CBL1 and CBL10 at the plasma membrane and the tonoplast, respectively. We have also visualized by mcBiFC the simultaneous formation of CBL1/CIPK1 and CBL9/CIPK1 protein complexes at the plasma membrane. Thus, mcBiFC provides a useful new tool to explore complex regulatory networks in plants.}, address = {Institut fr Botanik und Botanischer Garten, Universitt Mnster, Schlossplatz 4, 48149 Mnster, Germany; Max-Planck-Institut fr Molekulare Pflanzenphysiologie, Am Mhlenberg 1, 14476 Potsdam-Golm, Germany}, author = {Waadt, Rainer and Schmidt, Lena K. and Lohse, Marc and Hashimoto, Kenji and Bock, Ralph and Kudla, J\"{o}rg}, citeulike-article-id = {3055858}, citeulike-linkout-0 = {http://dx.doi.org/10.1111/j.1365-313X.2008.03612.x}, citeulike-linkout-1 = {http://www3.interscience.wiley.com/cgi-bin/abstract/120696360/ABSTRACT}, doi = {10.1111/j.1365-313X.2008.03612.x}, journal = {The Plant Journal}, keywords = {bifc, method}, number = {999A}, posted-at = {2008-07-29 09:39:35}, priority = {2}, title = {Multicolor bimolecular fluorescence complementation (mcBiFC) reveals simultaneous formation of alternative CBL/CIPK complexes in planta}, url = {http://dx.doi.org/10.1111/j.1365-313X.2008.03612.x}, volume = {9999}, year = {2008} }

TY - JOUR ID - citeulike:3055858 L3 - citeulike-article-id:3055858 N2 - The specificity of intracellular signaling and developmental patterning in biological systems relies on selective interactions between different proteins in specific cellular compartments. The identification of such protein-protein interactions is essential for unraveling complex signaling and regulatory networks. Recently, bimolecular fluorescence complementation (BiFC) has emerged as a powerful technique for the efficient detection of protein interactions in their native subcellular localization. Here we report significant technical advances in plant BiFC methodology. We describe a series of versatile BiFC vector sets that are fully compatible to previously generated vectors (Walter et al., 2004). The new vectors enable the generation of both C-terminal and N-terminal fusion proteins and carry optimized fluorescent protein genes that considerably improve the sensitivity of BiFC. Using these vectors, we describe a multicolor BiFC (mcBiFC) approach for the simultaneous visualization of multiple protein interactions in the same cell. Application to a protein interaction network acting in calcium-mediated signal transduction revealed the concurrent interaction of the protein kinase CIPK24 with the calcium sensors CBL1 and CBL10 at the plasma membrane and the tonoplast, respectively. We have also visualized by mcBiFC the simultaneous formation of CBL1/CIPK1 and CBL9/CIPK1 protein complexes at the plasma membrane. Thus, mcBiFC provides a useful new tool to explore complex regulatory networks in plants. IS - 999A JF - The Plant Journal AD - Institut fr Botanik und Botanischer Garten, Universitt Mnster, Schlossplatz 4, 48149 Mnster, Germany; Max-Planck-Institut fr Molekulare Pflanzenphysiologie, Am Mhlenberg 1, 14476 Potsdam-Golm, Germany TI - Multicolor bimolecular fluorescence complementation (mcBiFC) reveals simultaneous formation of alternative CBL/CIPK complexes in planta VL - 9999 KW - bifc KW - method AU - Waadt, Rainer AU - Schmidt, Lena AU - Lohse, Marc AU - Hashimoto, Kenji AU - Bock, Ralph AU - Kudla, Jörg PY - 2008/// UR - http://dx.doi.org/10.1111/j.1365-313X.2008.03612.x ER -