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Light plays an essential role in intracellular distribution of auxin efflux carrier PIN2 in Arabidopsis thaliana.

Laxmi A, Pan J, Morsy M, Chen R - PLoS ONE (2008)

Bottom Line: Using laser confocal scanning microscopy, and physiological and molecular genetic approaches, here, we show that in dark-grown seedlings, the PM localization of auxin efflux carrier PIN2 was largely reduced, and, in addition, PIN2 signal was detected in vacuolar compartments.In addition, the ubiquitin 26S proteasome is involved in the process, since its inhibition by mutations in COP9 and a proteasome inhibitor MG132 impaired the process.Based on these results, we postulate that light regulation of root development is mediated at least in part by changes in the intracellular distribution of auxin efflux carriers, PIN proteins, in response to the light environment.

View Article: PubMed Central - PubMed

Affiliation: Plant Biology Division, Samuel Roberts Noble Foundation, Ardmore, Oklahoma, USA.

ABSTRACT

Background: Light plays a key role in multiple plant developmental processes. It has been shown that root development is modulated by shoot-localized light signaling and requires shoot-derived transport of the plant hormone, auxin. However, the mechanism by which light regulates root development is not largely understood. In plants, the endogenous auxin, indole-3-acetic acid, is directionally transported by plasma-membrane (PM)-localized auxin influx and efflux carriers in transporting cells. Remarkably, the auxin efflux carrier PIN proteins exhibit asymmetric PM localization, determining the polarity of auxin transport. Similar to PM-resident receptors and transporters in animal and yeast cells, PIN proteins undergo constitutive cycling between the PM and endosomal compartments. Auxin plays multiple roles in PIN protein intracellular trafficking, inhibiting PIN2 endocytosis at some concentrations and promoting PIN2 degradation at others. However, how PIN proteins are turned over in plant cells is yet to be addressed.

Methodology and principle findings: Using laser confocal scanning microscopy, and physiological and molecular genetic approaches, here, we show that in dark-grown seedlings, the PM localization of auxin efflux carrier PIN2 was largely reduced, and, in addition, PIN2 signal was detected in vacuolar compartments. This is in contrast to light-grown seedlings where PIN2 was predominantly PM-localized. In light-grown plants after shift to dark or to continuous red or far-red light, PIN2 also accumulated in vacuolar compartments. We show that PIN2 vacuolar targeting was derived from the PM via endocytic trafficking and inhibited by HY5-dependent light signaling. In addition, the ubiquitin 26S proteasome is involved in the process, since its inhibition by mutations in COP9 and a proteasome inhibitor MG132 impaired the process.

Conclusions and significance: Collectively, our data indicate that light plays an essential role in PIN2 intracellular trafficking, promoting PM-localization in the presence of light and, on the other hand, vacuolar targeting for protein degradation in the absence of light. Based on these results, we postulate that light regulation of root development is mediated at least in part by changes in the intracellular distribution of auxin efflux carriers, PIN proteins, in response to the light environment.

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Localization of AUX1-YFP, PIP2A-GFP and deltaTIP-GFP in light- and dark-grown plants.AUX1-YFP was mainly localized to the basal PM of lateral root cap and central pre-vascular cells, and to the PM of root columella cells in both light-grown (A) and dark-grown (B) seedling roots. PIP2A-GFP was predominantly on the PM of all root cells, except that it was excluded from the quiescent center and surrounding initial cells, of light-grown plants (C). In dark-grown seedlings, a detectable level of PIP2A-GFP accumulated in vacuolar compartments (D). deltaTIP-GFP labeled both the PM and tonoplast membrane of root cells of light-grown (E) and dark-grown (F) plants. Shown in right and insets were close-up images. Scale bars, 50 µm (left), 25 µm (right).
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pone-0001510-g003: Localization of AUX1-YFP, PIP2A-GFP and deltaTIP-GFP in light- and dark-grown plants.AUX1-YFP was mainly localized to the basal PM of lateral root cap and central pre-vascular cells, and to the PM of root columella cells in both light-grown (A) and dark-grown (B) seedling roots. PIP2A-GFP was predominantly on the PM of all root cells, except that it was excluded from the quiescent center and surrounding initial cells, of light-grown plants (C). In dark-grown seedlings, a detectable level of PIP2A-GFP accumulated in vacuolar compartments (D). deltaTIP-GFP labeled both the PM and tonoplast membrane of root cells of light-grown (E) and dark-grown (F) plants. Shown in right and insets were close-up images. Scale bars, 50 µm (left), 25 µm (right).

Mentions: To determine whether the vacuolar accumulation of PIN2 occurred via a specific or general process, we examined localization patterns of several other PM-resident proteins. We observed that several other PIN proteins including PIN1 [32], [55] and PIN7 [36] similarly accumulated in vacuoles of several different types of root cells of dark-grown seedlings, where PIN proteins were expressed (Figs. 1H–K). Furthermore, a PM-localized water channel PIP2A [56] also changed from predominant PM-location in light-grown seedlings to both PM and vacuolar locations in dark-grown seedlings (Figs. 3C, D; insets). Surprisingly, the presumptive auxin influx carrier AUX1 did not significantly alter its intracellular localization in three different types of cells, root columella, lateral root cap and central pre-vascular cells of dark-grown seedlings (Figs. 3A, B; insets). The vacuolar structure, as indicated by the pattern of a deltaTIP-GFP fusion protein that marks both the PM and tonoplast membrane [56], [57], remained largely unchanged in the meristematic region of the root grown in dark compared to the light-grown counterpart (Figs. 3E, F; insets). Taken together, our data suggest that PIN2 vacuolar accumulation in roots of dark-grown seedlings takes place via a process that is shared by a subset of PM-resident proteins.


Light plays an essential role in intracellular distribution of auxin efflux carrier PIN2 in Arabidopsis thaliana.

Laxmi A, Pan J, Morsy M, Chen R - PLoS ONE (2008)

Localization of AUX1-YFP, PIP2A-GFP and deltaTIP-GFP in light- and dark-grown plants.AUX1-YFP was mainly localized to the basal PM of lateral root cap and central pre-vascular cells, and to the PM of root columella cells in both light-grown (A) and dark-grown (B) seedling roots. PIP2A-GFP was predominantly on the PM of all root cells, except that it was excluded from the quiescent center and surrounding initial cells, of light-grown plants (C). In dark-grown seedlings, a detectable level of PIP2A-GFP accumulated in vacuolar compartments (D). deltaTIP-GFP labeled both the PM and tonoplast membrane of root cells of light-grown (E) and dark-grown (F) plants. Shown in right and insets were close-up images. Scale bars, 50 µm (left), 25 µm (right).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2200863&req=5

pone-0001510-g003: Localization of AUX1-YFP, PIP2A-GFP and deltaTIP-GFP in light- and dark-grown plants.AUX1-YFP was mainly localized to the basal PM of lateral root cap and central pre-vascular cells, and to the PM of root columella cells in both light-grown (A) and dark-grown (B) seedling roots. PIP2A-GFP was predominantly on the PM of all root cells, except that it was excluded from the quiescent center and surrounding initial cells, of light-grown plants (C). In dark-grown seedlings, a detectable level of PIP2A-GFP accumulated in vacuolar compartments (D). deltaTIP-GFP labeled both the PM and tonoplast membrane of root cells of light-grown (E) and dark-grown (F) plants. Shown in right and insets were close-up images. Scale bars, 50 µm (left), 25 µm (right).
Mentions: To determine whether the vacuolar accumulation of PIN2 occurred via a specific or general process, we examined localization patterns of several other PM-resident proteins. We observed that several other PIN proteins including PIN1 [32], [55] and PIN7 [36] similarly accumulated in vacuoles of several different types of root cells of dark-grown seedlings, where PIN proteins were expressed (Figs. 1H–K). Furthermore, a PM-localized water channel PIP2A [56] also changed from predominant PM-location in light-grown seedlings to both PM and vacuolar locations in dark-grown seedlings (Figs. 3C, D; insets). Surprisingly, the presumptive auxin influx carrier AUX1 did not significantly alter its intracellular localization in three different types of cells, root columella, lateral root cap and central pre-vascular cells of dark-grown seedlings (Figs. 3A, B; insets). The vacuolar structure, as indicated by the pattern of a deltaTIP-GFP fusion protein that marks both the PM and tonoplast membrane [56], [57], remained largely unchanged in the meristematic region of the root grown in dark compared to the light-grown counterpart (Figs. 3E, F; insets). Taken together, our data suggest that PIN2 vacuolar accumulation in roots of dark-grown seedlings takes place via a process that is shared by a subset of PM-resident proteins.

Bottom Line: Using laser confocal scanning microscopy, and physiological and molecular genetic approaches, here, we show that in dark-grown seedlings, the PM localization of auxin efflux carrier PIN2 was largely reduced, and, in addition, PIN2 signal was detected in vacuolar compartments.In addition, the ubiquitin 26S proteasome is involved in the process, since its inhibition by mutations in COP9 and a proteasome inhibitor MG132 impaired the process.Based on these results, we postulate that light regulation of root development is mediated at least in part by changes in the intracellular distribution of auxin efflux carriers, PIN proteins, in response to the light environment.

View Article: PubMed Central - PubMed

Affiliation: Plant Biology Division, Samuel Roberts Noble Foundation, Ardmore, Oklahoma, USA.

ABSTRACT

Background: Light plays a key role in multiple plant developmental processes. It has been shown that root development is modulated by shoot-localized light signaling and requires shoot-derived transport of the plant hormone, auxin. However, the mechanism by which light regulates root development is not largely understood. In plants, the endogenous auxin, indole-3-acetic acid, is directionally transported by plasma-membrane (PM)-localized auxin influx and efflux carriers in transporting cells. Remarkably, the auxin efflux carrier PIN proteins exhibit asymmetric PM localization, determining the polarity of auxin transport. Similar to PM-resident receptors and transporters in animal and yeast cells, PIN proteins undergo constitutive cycling between the PM and endosomal compartments. Auxin plays multiple roles in PIN protein intracellular trafficking, inhibiting PIN2 endocytosis at some concentrations and promoting PIN2 degradation at others. However, how PIN proteins are turned over in plant cells is yet to be addressed.

Methodology and principle findings: Using laser confocal scanning microscopy, and physiological and molecular genetic approaches, here, we show that in dark-grown seedlings, the PM localization of auxin efflux carrier PIN2 was largely reduced, and, in addition, PIN2 signal was detected in vacuolar compartments. This is in contrast to light-grown seedlings where PIN2 was predominantly PM-localized. In light-grown plants after shift to dark or to continuous red or far-red light, PIN2 also accumulated in vacuolar compartments. We show that PIN2 vacuolar targeting was derived from the PM via endocytic trafficking and inhibited by HY5-dependent light signaling. In addition, the ubiquitin 26S proteasome is involved in the process, since its inhibition by mutations in COP9 and a proteasome inhibitor MG132 impaired the process.

Conclusions and significance: Collectively, our data indicate that light plays an essential role in PIN2 intracellular trafficking, promoting PM-localization in the presence of light and, on the other hand, vacuolar targeting for protein degradation in the absence of light. Based on these results, we postulate that light regulation of root development is mediated at least in part by changes in the intracellular distribution of auxin efflux carriers, PIN proteins, in response to the light environment.

Show MeSH
Related in: MedlinePlus