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Auxin regulates SNARE-dependent vacuolar morphology restricting cell size.

Löfke C, Dünser K, Scheuring D, Kleine-Vehn J - Elife (2015)

Bottom Line: Here, we reveal that the phytohormone auxin impacts on the shape of the biggest plant organelle, the vacuole.Genetic and pharmacological interference with the auxin effect on vacuolar SNAREs interrelates with auxin-resistant vacuolar morphogenesis and cell size regulation.Vacuolar SNARE VTI11 is strictly required for auxin-reliant vacuolar morphogenesis and loss of function renders cells largely insensitive to auxin-dependent growth inhibition.

View Article: PubMed Central - PubMed

Affiliation: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, Vienna, Austria.

ABSTRACT
The control of cellular growth is central to multicellular patterning. In plants, the encapsulating cell wall literally binds neighbouring cells to each other and limits cellular sliding/migration. In contrast to its developmental importance, growth regulation is poorly understood in plants. Here, we reveal that the phytohormone auxin impacts on the shape of the biggest plant organelle, the vacuole. TIR1/AFBs-dependent auxin signalling posttranslationally controls the protein abundance of vacuolar SNARE components. Genetic and pharmacological interference with the auxin effect on vacuolar SNAREs interrelates with auxin-resistant vacuolar morphogenesis and cell size regulation. Vacuolar SNARE VTI11 is strictly required for auxin-reliant vacuolar morphogenesis and loss of function renders cells largely insensitive to auxin-dependent growth inhibition. Our data suggests that the adaptation of SNARE-dependent vacuolar morphogenesis allows auxin to limit cellular expansion, contributing to root organ growth rates.

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Increase in SNARE intensity is independent of membrane crowding.(A–J) Simultaneous imaging of VAMP711-RFP/YFP and tonoplast staining dyes under untreated and high auxin conditions. pUBQ10::VAMP711-RFP (A and B) and pUBQ10::VAMP711-YFP (F and G) expressing seedlings were counterstained either with MDY-64 (C and D) or FM4-64 (H and I) and treated with DMSO (A and C) or NAA (B and D) (500 nM; 20 hr). (E) Quantification of mean grey value of VAMP711-RFP and MDY-64 after NAA treatments (500 nM) compared to DMSO control. (J) Quantification of mean grey value of VAMP711-YFP and FM4-64 after NAA treatments (500 nM) compared to DMSO control. Error bars represent s.e.m. For statistical analysis DMSO and NAA treatments were compared. n = 40 cells in 10 individual seedlings. Student's t-test p-values: ***p < 0.001. Scale bar: 15 µm.DOI:http://dx.doi.org/10.7554/eLife.05868.010
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fig4s1: Increase in SNARE intensity is independent of membrane crowding.(A–J) Simultaneous imaging of VAMP711-RFP/YFP and tonoplast staining dyes under untreated and high auxin conditions. pUBQ10::VAMP711-RFP (A and B) and pUBQ10::VAMP711-YFP (F and G) expressing seedlings were counterstained either with MDY-64 (C and D) or FM4-64 (H and I) and treated with DMSO (A and C) or NAA (B and D) (500 nM; 20 hr). (E) Quantification of mean grey value of VAMP711-RFP and MDY-64 after NAA treatments (500 nM) compared to DMSO control. (J) Quantification of mean grey value of VAMP711-YFP and FM4-64 after NAA treatments (500 nM) compared to DMSO control. Error bars represent s.e.m. For statistical analysis DMSO and NAA treatments were compared. n = 40 cells in 10 individual seedlings. Student's t-test p-values: ***p < 0.001. Scale bar: 15 µm.DOI:http://dx.doi.org/10.7554/eLife.05868.010

Mentions: In the following we got interested in SNAP (Soluble NSF Attachment Protein) Receptor (SNARE) complexes at the vacuole. Proximity of adjacent membrane allows the interaction of v (vesicle)- and t (target)-SNAREs to form a complex, allowing the fusion of vesicles to specific target membranes. SNAREs are essential for eukaryotic vesicle trafficking and according to structural features SNAREs are divided in R (arginine)- and Q (glutamine)-SNAREs (Martens and McMahon, 2008). In yeast, the SNARE complex is furthermore central in homotypic vacuolar membrane remodelling and proteomic approaches have identified conserved SNARE complexes at the plant tonoplast (Carter et al., 2004). Ergo, we tested whether auxin affects vacuolar SNAREs in Arabidopsis. Remarkably, increased auxin biosynthesis or exogenous application of auxin increased the fluorescence intensity of tonoplast localised SNAREs, such as VAMP711-YFP, SYP21-YFP and SYP22-GFP (Figure 4A–L,N,O). Auxin severely impacts on vacuolar appearance and, hence, it could be that membrane crowding induces higher fluorescence. To address this question we performed co-localisation of VAMP711-RFP/YFP and membrane dyes, such as FM4-64 and MDY-64. Notably, VAMP711-RFP/YFP, but not the membrane dyes showed auxin-induced signal intensities, suggesting that the auxin effect on vacuolar SNAREs does not rely on membrane crowding (Figure 4—figure supplement 1).10.7554/eLife.05868.009Figure 4.Auxin posttranslationally stabilises tonoplast localised SNAREs.


Auxin regulates SNARE-dependent vacuolar morphology restricting cell size.

Löfke C, Dünser K, Scheuring D, Kleine-Vehn J - Elife (2015)

Increase in SNARE intensity is independent of membrane crowding.(A–J) Simultaneous imaging of VAMP711-RFP/YFP and tonoplast staining dyes under untreated and high auxin conditions. pUBQ10::VAMP711-RFP (A and B) and pUBQ10::VAMP711-YFP (F and G) expressing seedlings were counterstained either with MDY-64 (C and D) or FM4-64 (H and I) and treated with DMSO (A and C) or NAA (B and D) (500 nM; 20 hr). (E) Quantification of mean grey value of VAMP711-RFP and MDY-64 after NAA treatments (500 nM) compared to DMSO control. (J) Quantification of mean grey value of VAMP711-YFP and FM4-64 after NAA treatments (500 nM) compared to DMSO control. Error bars represent s.e.m. For statistical analysis DMSO and NAA treatments were compared. n = 40 cells in 10 individual seedlings. Student's t-test p-values: ***p < 0.001. Scale bar: 15 µm.DOI:http://dx.doi.org/10.7554/eLife.05868.010
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fig4s1: Increase in SNARE intensity is independent of membrane crowding.(A–J) Simultaneous imaging of VAMP711-RFP/YFP and tonoplast staining dyes under untreated and high auxin conditions. pUBQ10::VAMP711-RFP (A and B) and pUBQ10::VAMP711-YFP (F and G) expressing seedlings were counterstained either with MDY-64 (C and D) or FM4-64 (H and I) and treated with DMSO (A and C) or NAA (B and D) (500 nM; 20 hr). (E) Quantification of mean grey value of VAMP711-RFP and MDY-64 after NAA treatments (500 nM) compared to DMSO control. (J) Quantification of mean grey value of VAMP711-YFP and FM4-64 after NAA treatments (500 nM) compared to DMSO control. Error bars represent s.e.m. For statistical analysis DMSO and NAA treatments were compared. n = 40 cells in 10 individual seedlings. Student's t-test p-values: ***p < 0.001. Scale bar: 15 µm.DOI:http://dx.doi.org/10.7554/eLife.05868.010
Mentions: In the following we got interested in SNAP (Soluble NSF Attachment Protein) Receptor (SNARE) complexes at the vacuole. Proximity of adjacent membrane allows the interaction of v (vesicle)- and t (target)-SNAREs to form a complex, allowing the fusion of vesicles to specific target membranes. SNAREs are essential for eukaryotic vesicle trafficking and according to structural features SNAREs are divided in R (arginine)- and Q (glutamine)-SNAREs (Martens and McMahon, 2008). In yeast, the SNARE complex is furthermore central in homotypic vacuolar membrane remodelling and proteomic approaches have identified conserved SNARE complexes at the plant tonoplast (Carter et al., 2004). Ergo, we tested whether auxin affects vacuolar SNAREs in Arabidopsis. Remarkably, increased auxin biosynthesis or exogenous application of auxin increased the fluorescence intensity of tonoplast localised SNAREs, such as VAMP711-YFP, SYP21-YFP and SYP22-GFP (Figure 4A–L,N,O). Auxin severely impacts on vacuolar appearance and, hence, it could be that membrane crowding induces higher fluorescence. To address this question we performed co-localisation of VAMP711-RFP/YFP and membrane dyes, such as FM4-64 and MDY-64. Notably, VAMP711-RFP/YFP, but not the membrane dyes showed auxin-induced signal intensities, suggesting that the auxin effect on vacuolar SNAREs does not rely on membrane crowding (Figure 4—figure supplement 1).10.7554/eLife.05868.009Figure 4.Auxin posttranslationally stabilises tonoplast localised SNAREs.

Bottom Line: Here, we reveal that the phytohormone auxin impacts on the shape of the biggest plant organelle, the vacuole.Genetic and pharmacological interference with the auxin effect on vacuolar SNAREs interrelates with auxin-resistant vacuolar morphogenesis and cell size regulation.Vacuolar SNARE VTI11 is strictly required for auxin-reliant vacuolar morphogenesis and loss of function renders cells largely insensitive to auxin-dependent growth inhibition.

View Article: PubMed Central - PubMed

Affiliation: Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, Vienna, Austria.

ABSTRACT
The control of cellular growth is central to multicellular patterning. In plants, the encapsulating cell wall literally binds neighbouring cells to each other and limits cellular sliding/migration. In contrast to its developmental importance, growth regulation is poorly understood in plants. Here, we reveal that the phytohormone auxin impacts on the shape of the biggest plant organelle, the vacuole. TIR1/AFBs-dependent auxin signalling posttranslationally controls the protein abundance of vacuolar SNARE components. Genetic and pharmacological interference with the auxin effect on vacuolar SNAREs interrelates with auxin-resistant vacuolar morphogenesis and cell size regulation. Vacuolar SNARE VTI11 is strictly required for auxin-reliant vacuolar morphogenesis and loss of function renders cells largely insensitive to auxin-dependent growth inhibition. Our data suggests that the adaptation of SNARE-dependent vacuolar morphogenesis allows auxin to limit cellular expansion, contributing to root organ growth rates.

Show MeSH
Related in: MedlinePlus