Combined Bimolecular Fluorescence Complementation and Förster Resonance Energy Transfer Reveals Ternary SNARE Complex Formation in Living Plant Cells

Kwaaitaal, Mark; Keinath, Nana F.; Pajonk, Simone; Biskup, Christoph; Panstruga, Ralph

doi:10.1104/pp.109.151142

Cited by 63 publications

(47 citation statements)

References 54 publications

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“…In barley (Hordeum vulgare), PIP2 colocalized in ROR2/SYP121 vesicle-like compartments, supporting this hypothesis (Kwaaitaal et al, 2010). We report here that the SYP121-Sp2 from Arabidopsis and its putative ortholog from maize affect the delivery of Zm-PIP2;5 to the plasma membrane in maize mesophyll protoplasts and epidermal cells.…”

Section: Introductionsupporting

confidence: 69%

“…For the photobleaching method, three images were acquired for both the mCFP and mYFP channels, and then the mYFP was bleached by 80 iterations with fullpower laser illumination, while the fluorescence intensity of the mCFP was simultaneously monitored along time. The FRET efficiency (E) was determined according the equation E = 12 I DA /I D , where I DA is the intensity of the donor fluorescence before photobleaching and I D the fluorescence intensity of the donor after photobleaching, as described elsewhere (Kwaaitaal et al, 2010). The sensitized emission method was used on a subpopulation of tobacco epidermal cells and protoplasts to confirm the data obtained by acceptor photobleaching.…”

Section: Confocal Microscopymentioning

confidence: 99%

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Selective Regulation of Maize Plasma Membrane Aquaporin Trafficking and Activity by the SNARE SYP121

et al. 2012

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Plasma membrane intrinsic proteins (PIPs) are aquaporins facilitating the diffusion of water through the cell membrane. We previously showed that the traffic of the maize (Zea mays) PIP2;5 to the plasma membrane is dependent on the endoplasmic reticulum diacidic export motif. Here, we report that the post-Golgi traffic and water channel activity of PIP2;5 are regulated by the SNARE (for soluble N-ethylmaleimide-sensitive factor protein attachment protein receptor) SYP121, a plasma membrane resident syntaxin involved in vesicle traffic, signaling, and regulation of K + channels. We demonstrate that the expression of the dominant-negative SYP121-Sp2 fragment in maize mesophyll protoplasts or epidermal cells leads to a decrease in the delivery of PIP2;5 to the plasma membrane. Protoplast and oocyte swelling assays showed that PIP2;5 water channel activity is negatively affected by SYP121-Sp2. A combination of in vitro (copurification assays) and in vivo (bimolecular fluorescence complementation, Förster resonance energy transfer, and yeast split-ubiquitin) approaches allowed us to demonstrate that SYP121 and PIP2;5 physically interact. Together with previous data demonstrating the role of SYP121 in regulating K + channel trafficking and activity, these results suggest that SYP121 SNARE contributes to the regulation of the cell osmotic homeostasis.

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Section: Introductionsupporting

confidence: 69%

Section: Confocal Microscopymentioning

confidence: 99%

Selective Regulation of Maize Plasma Membrane Aquaporin Trafficking and Activity by the SNARE SYP121

et al. 2012

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“…SYP61 has thus far been considered solely as a TGN resident, with no involvement in anterograde transport to the PM. It has been shown that the SYP121 Qa-SNARE PM resident (PEN1) [51] interacts with the SNAP33 adaptor and the two functionally redundant VAMP72 synaptobrevins [52] in forming the PEN1-SNAP33-VAMP721/VAMP722 tertiary SNARE complex that contributes to a fungal resistance system [52,53]. In the leaf epidermis, VAMP722-GFP vesicles accumulate in intracellular compartments in response to an infection by Blumeria graminis.…”

Section: Syp61 Facilitates the Trafficking Of Proteins To The Pmmentioning

confidence: 99%

Isolation and proteomic analysis of the SYP61 compartment reveal its role in exocytic trafficking in Arabidopsis

et al. 2011

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The endomembrane system is a complex and dynamic intracellular trafficking network. It is very challenging to track individual vesicles and their cargos in real time; however, affinity purification allows vesicles to be isolated in their natural state so that their constituent proteins can be identified. Pioneering this approach in plants, we isolated the SYP61 trans-Golgi network compartment and carried out a comprehensive proteomic analysis of its contents with only minimal interference from other organelles. The proteome of SYP61 revealed the association of proteins of unknown function that have previously not been ascribed to this compartment. We identified a complete SYP61 SNARE complex, including regulatory proteins and validated the proteome data by showing that several of these proteins associated with SYP61 in planta. We further identified the SYP121-complex and cellulose synthases, suggesting that SYP61 plays a role in the exocytic trafficking and the transport of cell wall components to the plasma membrane. The presence of proteins of unknown function in the SYP61 proteome including ECHIDNA offers the opportunity to identify novel trafficking components and cargos. The affinity purification of plant vesicles in their natural state provides a basis for further analysis and dissection of complex endomembrane networks. The approach is widely applicable and can afford the study of several vesicle populations in plants, which can be compared with the SYP61 vesicle proteome.

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“…Additionally, the CFP-HDA19 and YFP-SNL1 fusion vectors were constructed to confirm the fusion protein localization with pENSG-CFP and pENSG-YFP, respectively (Kwaaitaal et al, 2010). For transient expression, Agrobacterium strains (GV3101 pm90RK) carrying the constructs were used (Clough and Bent, 1998) for infiltration of 5-to 6-week-old Nicotiana benthamiana leaves.…”

Section: Interaction Between Snl1 and Hda19 By Plant Bifcmentioning

confidence: 99%

ArabidopsisPaired Amphipathic Helix Proteins SNL1 and SNL2 Redundantly Regulate Primary Seed Dormancy via Abscisic Acid–Ethylene Antagonism Mediated by Histone Deacetylation

et al. 2013

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Histone (de)acetylation is a highly conserved chromatin modification that is vital for development and growth. In this study, we identified a role in seed dormancy for two members of the histone deacetylation complex in Arabidopsis thaliana, SIN3-LIKE1 (SNL1) and SNL2. The double mutant snl1 snl2 shows reduced dormancy and hypersensitivity to the histone deacetylase inhibitors trichostatin A and diallyl disulfide compared with the wild type. SNL1 interacts with HISTONE DEACETYLASE19 in vitro and in planta, and loss-of-function mutants of SNL1 and SNL2 show increased acetylation levels of histone 3 lysine 9/18 (H3K9/18) and H3K14. Moreover, SNL1 and SNL2 regulate key genes involved in the ethylene and abscisic acid (ABA) pathways by decreasing their histone acetylation levels. Taken together, we showed that SNL1 and SNL2 regulate seed dormancy by mediating the ABA-ethylene antagonism in Arabidopsis. SNL1 and SNL2 could represent a crosslink point of the ABA and ethylene pathways in the regulation of seed dormancy.

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Combined Bimolecular Fluorescence Complementation and Förster Resonance Energy Transfer Reveals Ternary SNARE Complex Formation in Living Plant Cells

Cited by 63 publications

References 54 publications

Selective Regulation of Maize Plasma Membrane Aquaporin Trafficking and Activity by the SNARE SYP121

Selective Regulation of Maize Plasma Membrane Aquaporin Trafficking and Activity by the SNARE SYP121

Isolation and proteomic analysis of the SYP61 compartment reveal its role in exocytic trafficking in Arabidopsis

ArabidopsisPaired Amphipathic Helix Proteins SNL1 and SNL2 Redundantly Regulate Primary Seed Dormancy via Abscisic Acid–Ethylene Antagonism Mediated by Histone Deacetylation

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