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Physiological and Molecular Responses to Excess Boron in Citrus macrophylla W.

Martínez-Cuenca MR, Martínez-Alcántara B, Quiñones A, Ruiz M, Iglesias DJ, Primo-Millo E, Forner-Giner MÁ - PLoS ONE (2015)

Bottom Line: Excess B led to high B concentration in +B plants (3.8- and 1.4-fold in leaves and roots, respectively) when compared with Ct ones.Finally, excess B caused a significant rise in proline concentration (51% and 34% in roots and leaves, respectively), while the MDA level did not exceed 20%.In conclusion, Cm tolerance to a high B level is likely based on the synergism of several specific mechanisms against B toxicity, including: 1/ down-regulation of NIP5 and PIP1 boron transporters; 2/ activation of B efflux from cells due to the up-regulation of putative BOR4 gene; 3/ compartmentation of B in the vacuole through TIP5 transporter activation and the acidification of the organelle; 4/ insolubilisation of B and deposition in cell walls preventing from cytoplasm damage; and, 5/ induction of an efficient antioxidant system through proline accumulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.

ABSTRACT
This work provides insight into several mechanisms involved in boron (B) regulation pathway in response to high B conditions in Citrus. The study was carried out in Citrus macrophylla W. (Cm) seedlings cultured "in vitro" in media with 50 or 400 μM H3BO3 (control, Ct, and B-excess, +B, plants, respectively). Growth parameters, B concentration, leaf chlorophyll (Chl) concentration, the expression of the main putative genes involved in B transport and distribution, and leaf and root proline and malonaldehyde (MDA) concentrations, were assessed. Excess B led to high B concentration in +B plants (3.8- and 1.4-fold in leaves and roots, respectively) when compared with Ct ones. However, a minor effect was recorded in the plant (incipient visual symptoms, less than 27% reduction in root growth and 26% decrease in Chl b concentration). B toxicity down-regulated by half the expression level of putative B transporter genes NIP5 and PIP1. CmBOR1 gene was not repressed in +B plants and B accumulated in the shoots. High B level increased the transcripts of putative gene TIP5, involved in B transport across the tonoplast, by 3.3- and 2.4-fold in leaves and roots, respectively. The activity of V-PPiase proton pump, related with the electrochemical gradient in the vacuole, was also enhanced in +B organs. B toxicity up-regulated putative BOR4 gene (2.1- and 2.7-fold in roots and leaves, respectively), which codifies for an active efflux B transporter. Accordingly, B was located in +B plants preferently in an insoluble form on cell walls. Finally, excess B caused a significant rise in proline concentration (51% and 34% in roots and leaves, respectively), while the MDA level did not exceed 20%. In conclusion, Cm tolerance to a high B level is likely based on the synergism of several specific mechanisms against B toxicity, including: 1/ down-regulation of NIP5 and PIP1 boron transporters; 2/ activation of B efflux from cells due to the up-regulation of putative BOR4 gene; 3/ compartmentation of B in the vacuole through TIP5 transporter activation and the acidification of the organelle; 4/ insolubilisation of B and deposition in cell walls preventing from cytoplasm damage; and, 5/ induction of an efficient antioxidant system through proline accumulation.

No MeSH data available.


Related in: MedlinePlus

Relative expression of NIP5, PIP1 and PIP2 genes measured by real-time RT-PCR analysis, in roots and leaves of Citrus macrophylla seedlings grown for 25 days in B-normal (50 μM, Ct) and B-toxic (400 μM, +B) nutrient solutions.Values are the means ± SE of three independent experiments (n = 3). For a comparison of means, an ANOVA followed by the LSD test, calculated at 95% confidence level, was performed. Different letters in the same organ indicate significant differences between treatments (P <0.05).
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pone.0134372.g003: Relative expression of NIP5, PIP1 and PIP2 genes measured by real-time RT-PCR analysis, in roots and leaves of Citrus macrophylla seedlings grown for 25 days in B-normal (50 μM, Ct) and B-toxic (400 μM, +B) nutrient solutions.Values are the means ± SE of three independent experiments (n = 3). For a comparison of means, an ANOVA followed by the LSD test, calculated at 95% confidence level, was performed. Different letters in the same organ indicate significant differences between treatments (P <0.05).

Mentions: Acquisition of boron by cells is associated with the activity of some members of the aquaporin family codified by the genes NIP5, PIP1 and PIP2 (Fig 3). The expression level of putative NIP5 gene lowered in roots as a result of high B levels. Thus, +B seedlings contained 52% fewer NIP5 transcripts in roots than Ct ones. It is noteworthy that no effect on the NIP5 transcript level was recorded when analysed in +B leaves. In A. thaliana, this gene codified a boric acid channel for efficient B acquisition under limited B supply [12]. This result was also found in Carrizo citrange roots, where CiNIP5 expression increased under B-deficiency conditions [31]. In contrast, its down-expression under B-excess conditions in the roots of Citrus plants has revealed a point of regulation of B uptake efficiency of different citrus rootstocks. Similar behaviour had been previously described by Schnurbusch et al. [39], which showed a relation between HvNIP2:1 gene expression and B toxicity tolerance in barley. Moreover, Tanaka et al. [40] suggested the importance of AtNIP5:1 degradation for plant acclimation to high B conditions.


Physiological and Molecular Responses to Excess Boron in Citrus macrophylla W.

Martínez-Cuenca MR, Martínez-Alcántara B, Quiñones A, Ruiz M, Iglesias DJ, Primo-Millo E, Forner-Giner MÁ - PLoS ONE (2015)

Relative expression of NIP5, PIP1 and PIP2 genes measured by real-time RT-PCR analysis, in roots and leaves of Citrus macrophylla seedlings grown for 25 days in B-normal (50 μM, Ct) and B-toxic (400 μM, +B) nutrient solutions.Values are the means ± SE of three independent experiments (n = 3). For a comparison of means, an ANOVA followed by the LSD test, calculated at 95% confidence level, was performed. Different letters in the same organ indicate significant differences between treatments (P <0.05).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4520451&req=5

pone.0134372.g003: Relative expression of NIP5, PIP1 and PIP2 genes measured by real-time RT-PCR analysis, in roots and leaves of Citrus macrophylla seedlings grown for 25 days in B-normal (50 μM, Ct) and B-toxic (400 μM, +B) nutrient solutions.Values are the means ± SE of three independent experiments (n = 3). For a comparison of means, an ANOVA followed by the LSD test, calculated at 95% confidence level, was performed. Different letters in the same organ indicate significant differences between treatments (P <0.05).
Mentions: Acquisition of boron by cells is associated with the activity of some members of the aquaporin family codified by the genes NIP5, PIP1 and PIP2 (Fig 3). The expression level of putative NIP5 gene lowered in roots as a result of high B levels. Thus, +B seedlings contained 52% fewer NIP5 transcripts in roots than Ct ones. It is noteworthy that no effect on the NIP5 transcript level was recorded when analysed in +B leaves. In A. thaliana, this gene codified a boric acid channel for efficient B acquisition under limited B supply [12]. This result was also found in Carrizo citrange roots, where CiNIP5 expression increased under B-deficiency conditions [31]. In contrast, its down-expression under B-excess conditions in the roots of Citrus plants has revealed a point of regulation of B uptake efficiency of different citrus rootstocks. Similar behaviour had been previously described by Schnurbusch et al. [39], which showed a relation between HvNIP2:1 gene expression and B toxicity tolerance in barley. Moreover, Tanaka et al. [40] suggested the importance of AtNIP5:1 degradation for plant acclimation to high B conditions.

Bottom Line: Excess B led to high B concentration in +B plants (3.8- and 1.4-fold in leaves and roots, respectively) when compared with Ct ones.Finally, excess B caused a significant rise in proline concentration (51% and 34% in roots and leaves, respectively), while the MDA level did not exceed 20%.In conclusion, Cm tolerance to a high B level is likely based on the synergism of several specific mechanisms against B toxicity, including: 1/ down-regulation of NIP5 and PIP1 boron transporters; 2/ activation of B efflux from cells due to the up-regulation of putative BOR4 gene; 3/ compartmentation of B in the vacuole through TIP5 transporter activation and the acidification of the organelle; 4/ insolubilisation of B and deposition in cell walls preventing from cytoplasm damage; and, 5/ induction of an efficient antioxidant system through proline accumulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.

ABSTRACT
This work provides insight into several mechanisms involved in boron (B) regulation pathway in response to high B conditions in Citrus. The study was carried out in Citrus macrophylla W. (Cm) seedlings cultured "in vitro" in media with 50 or 400 μM H3BO3 (control, Ct, and B-excess, +B, plants, respectively). Growth parameters, B concentration, leaf chlorophyll (Chl) concentration, the expression of the main putative genes involved in B transport and distribution, and leaf and root proline and malonaldehyde (MDA) concentrations, were assessed. Excess B led to high B concentration in +B plants (3.8- and 1.4-fold in leaves and roots, respectively) when compared with Ct ones. However, a minor effect was recorded in the plant (incipient visual symptoms, less than 27% reduction in root growth and 26% decrease in Chl b concentration). B toxicity down-regulated by half the expression level of putative B transporter genes NIP5 and PIP1. CmBOR1 gene was not repressed in +B plants and B accumulated in the shoots. High B level increased the transcripts of putative gene TIP5, involved in B transport across the tonoplast, by 3.3- and 2.4-fold in leaves and roots, respectively. The activity of V-PPiase proton pump, related with the electrochemical gradient in the vacuole, was also enhanced in +B organs. B toxicity up-regulated putative BOR4 gene (2.1- and 2.7-fold in roots and leaves, respectively), which codifies for an active efflux B transporter. Accordingly, B was located in +B plants preferently in an insoluble form on cell walls. Finally, excess B caused a significant rise in proline concentration (51% and 34% in roots and leaves, respectively), while the MDA level did not exceed 20%. In conclusion, Cm tolerance to a high B level is likely based on the synergism of several specific mechanisms against B toxicity, including: 1/ down-regulation of NIP5 and PIP1 boron transporters; 2/ activation of B efflux from cells due to the up-regulation of putative BOR4 gene; 3/ compartmentation of B in the vacuole through TIP5 transporter activation and the acidification of the organelle; 4/ insolubilisation of B and deposition in cell walls preventing from cytoplasm damage; and, 5/ induction of an efficient antioxidant system through proline accumulation.

No MeSH data available.


Related in: MedlinePlus