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Single-cell-based system to monitor carrier driven cellular auxin homeostasis

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Abstract
Background: Abundance and distribution of the plant hormone auxin play important roles in plant development. Besides other metabolic processes, various auxin carriers control the cellular level of active auxin and, hence, are major regulators of cellular auxin homeostasis. Despite the developmental importance of auxin transporters, a simple medium-to-high throughput approach to assess carrier activities is still missing. Here we show that carrier driven depletion of cellular auxin correlates with reduced nuclear auxin signaling in tobacco Bright Yellow-2 (BY-2) cell cultures. Results: We developed an easy to use transient single-cell-based system to detect carrier activity. We use the relative changes in signaling output of the auxin responsive promoter element DR5 to indirectly visualize auxin carrier activity. The feasibility of the transient approach was demonstrated by pharmacological and genetic interference with auxin signaling and transport. As a proof of concept, we provide visual evidence that the prominent auxin transport proteins PIN-FORMED (PIN) 2 and PIN5 regulate cellular auxin homeostasis at the plasma membrane and endoplasmic reticulum (ER), respectively. Our data suggest that PIN2 and PIN5 have different sensitivities to the auxin transport inhibitor 1-naphthylphthalamic acid (NPA). Also the putative PIN-LIKES (PILS) auxin carrier activity at the ER is insensitive to NPA in our system, indicating that NPA blocks intercellular, but not intracellular auxin transport. Conclusions: This single-cell-based system is a useful tool by which the activity of putative auxin carriers, such as PINs, PILS and WALLS ARE THIN1 (WAT1), can be indirectly visualized in a medium-to-high throughput manner. Moreover, our single cell system might be useful to investigate also other hormonal signaling pathways, such as cytokinin.
Keywords
TRANSPORT, ARABIDOPSIS, EFFLUX, TRANSFORMATION, PIN PROTEINS, AUX/IAA PROTEINS, PLANT-CELLS, PLASMA-MEMBRANE, ROOT HAIR-CELLS, CULTURED TOBACCO CELLS, Auxin transport, Auxin carrier, Auxin homeostasis, DR5

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Chicago
Barbez, Elke, Martina Laňková, Markéta Pařezová, Eva Zažímalová, Jan Petrášek, Jiri Friml, and Jürgen Kleine-Vehn. 2013. “Single-cell-based System to Monitor Carrier Driven Cellular Auxin Homeostasis.” Bmc Plant Biology 13.
APA
Barbez, E., Laňková, M., Pařezová, M., Zažímalová, E., Petrášek, J., Friml, J., & Kleine-Vehn, J. (2013). Single-cell-based system to monitor carrier driven cellular auxin homeostasis. BMC PLANT BIOLOGY, 13.
Vancouver
1.
Barbez E, Laňková M, Pařezová M, Zažímalová E, Petrášek J, Friml J, et al. Single-cell-based system to monitor carrier driven cellular auxin homeostasis. BMC PLANT BIOLOGY. 2013;13.
MLA
Barbez, Elke, Martina Laňková, Markéta Pařezová, et al. “Single-cell-based System to Monitor Carrier Driven Cellular Auxin Homeostasis.” BMC PLANT BIOLOGY 13 (2013): n. pag. Print.
@article{4196952,
  abstract     = {Background: Abundance and distribution of the plant hormone auxin play important roles in plant development. Besides other metabolic processes, various auxin carriers control the cellular level of active auxin and, hence, are major regulators of cellular auxin homeostasis. Despite the developmental importance of auxin transporters, a simple medium-to-high throughput approach to assess carrier activities is still missing. Here we show that carrier driven depletion of cellular auxin correlates with reduced nuclear auxin signaling in tobacco Bright Yellow-2 (BY-2) cell cultures. 
Results: We developed an easy to use transient single-cell-based system to detect carrier activity. We use the relative changes in signaling output of the auxin responsive promoter element DR5 to indirectly visualize auxin carrier activity. The feasibility of the transient approach was demonstrated by pharmacological and genetic interference with auxin signaling and transport. As a proof of concept, we provide visual evidence that the prominent auxin transport proteins PIN-FORMED (PIN) 2 and PIN5 regulate cellular auxin homeostasis at the plasma membrane and endoplasmic reticulum (ER), respectively. Our data suggest that PIN2 and PIN5 have different sensitivities to the auxin transport inhibitor 1-naphthylphthalamic acid (NPA). Also the putative PIN-LIKES (PILS) auxin carrier activity at the ER is insensitive to NPA in our system, indicating that NPA blocks intercellular, but not intracellular auxin transport. 
Conclusions: This single-cell-based system is a useful tool by which the activity of putative auxin carriers, such as PINs, PILS and WALLS ARE THIN1 (WAT1), can be indirectly visualized in a medium-to-high throughput manner. Moreover, our single cell system might be useful to investigate also other hormonal signaling pathways, such as cytokinin.},
  articleno    = {20},
  author       = {Barbez, Elke and La\v{n}kov{\'a}, Martina and Pa\v{r}ezov{\'a}, Mark{\'e}ta and Za\v{z}{\'i}malov{\'a}, Eva and Petr{\'a}\v{s}ek, Jan and Friml, Jiri and Kleine-Vehn, J{\"u}rgen},
  issn         = {1471-2229},
  journal      = {BMC PLANT BIOLOGY},
  language     = {eng},
  pages        = {13},
  title        = {Single-cell-based system to monitor carrier driven cellular auxin homeostasis},
  url          = {http://dx.doi.org/10.1186/1471-2229-13-20},
  volume       = {13},
  year         = {2013},
}

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