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Overexpression of a soybean ariadne-like ubiquitin ligase gene GmARI1 enhances aluminum tolerance in Arabidopsis.

Zhang X, Wang N, Chen P, Gao M, Liu J, Wang Y, Zhao T, Li Y, Gai J - PLoS ONE (2014)

Bottom Line: In vitro ubiquitination assay showed GmARI1 protein has E3 ligase activity.Overexpression of GmARI1 significantly enhanced the aluminum tolerance of transgenic Arabidopsis.These findings suggest that GmARI1 encodes a RBR type E3 ligase, which may play important roles in plant tolerance to aluminum stress.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Crop Genetics and Germplasm Enhancement, National Center for Soybean Improvement, Key Laboratory for Biology and Genetic Improvement of Soybean (General, Ministry of Agriculture), Nanjing Agricultural University, Nanjing, Jiangsu, China.

ABSTRACT
Ariadne (ARI) subfamily of RBR (Ring Between Ring fingers) proteins have been found as a group of putative E3 ubiquitin ligases containing RING (Really Interesting New Gene) finger domains in fruitfly, mouse, human and Arabidopsis. Recent studies showed several RING-type E3 ubiquitin ligases play important roles in plant response to abiotic stresses, but the function of ARI in plants is largely unknown. In this study, an ariadne-like E3 ubiquitin ligase gene was isolated from soybean, Glycine max (L.) Merr., and designated as GmARI1. It encodes a predicted protein of 586 amino acids with a RBR supra-domain. Subcellular localization studies using Arabidopsis protoplast cells indicated GmARI protein was located in nucleus. The expression of GmARI1 in soybean roots was induced as early as 2-4 h after simulated stress treatments such as aluminum, which coincided with the fact of aluminum toxicity firstly and mainly acting on plant roots. In vitro ubiquitination assay showed GmARI1 protein has E3 ligase activity. Overexpression of GmARI1 significantly enhanced the aluminum tolerance of transgenic Arabidopsis. These findings suggest that GmARI1 encodes a RBR type E3 ligase, which may play important roles in plant tolerance to aluminum stress.

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Performance of the transgenic Arabidopsis under 15 µM Al treatment.A. Root phenotypes of the 35S: GmARI1 overexpression lines GmARI1-3 and the wild type Col-0 grown on 15 µM Al in 1/2 MS (+Al, pH4.3), and 0 µM Al in 1/2 MS (-Al, pH4.3). B. Relative root growth (RRG, %) of the transgenic Arabidopsis lines (GmARI1-1, GmARI1-2, GmARI1-3) and the wild type (Col-0). RRG was calculated by the root growth length under Al treatment (15 µM Al in 1/2 MS, pH4.3) divided by the root growth length under control (0 µM Al in 1/2 MS, pH4.3). Error bars represent the standard error (SE), ** indicate the significance level of 0.01 by t-tests.
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pone-0111120-g009: Performance of the transgenic Arabidopsis under 15 µM Al treatment.A. Root phenotypes of the 35S: GmARI1 overexpression lines GmARI1-3 and the wild type Col-0 grown on 15 µM Al in 1/2 MS (+Al, pH4.3), and 0 µM Al in 1/2 MS (-Al, pH4.3). B. Relative root growth (RRG, %) of the transgenic Arabidopsis lines (GmARI1-1, GmARI1-2, GmARI1-3) and the wild type (Col-0). RRG was calculated by the root growth length under Al treatment (15 µM Al in 1/2 MS, pH4.3) divided by the root growth length under control (0 µM Al in 1/2 MS, pH4.3). Error bars represent the standard error (SE), ** indicate the significance level of 0.01 by t-tests.

Mentions: The gene expression of GmARI1 was induced early by Al stress, therefore we further studied its function in transgenic plants. An expression plasmid vector of pMDC83-GmARI1 was constructed and introduced into Arabidopsis plants using floral dip method. Transgenic T3 Arabidopsis over-expressing GmARI1 were generated and the positive transgenic lines were identified by RT-PCR (Fig. S2). Seeds of three T3 homozygous transgenic lines (GmARI1-1, GmARI1-2, and GmARI1-3) and wild type Col-0 were germinated on 1/2 MS medium. After 10 days, the seedlings were transferred to 1/2 MS (pH4.3) containing 15 µM Al or 0 µM Al as a control. Fifteen days later, the root growth of the wild type Col-0 was severely inhibited by 15 µM Al as compared with the control medium (0 µM), while the transgenic lines were little affected by Al (Fig. 9 A). The relative root growth (RRG) of the transgenic lines was significantly (p<0.01) longer than the wild type under Al treatment (Fig. 9B). The relative abundance of GmARI1 in the transgenic lines of GmARI1-1 and GmARI1-2 is higher than GmARI1-3, which coincided with RRG result (Fig. S3 and Fig. 9B).


Overexpression of a soybean ariadne-like ubiquitin ligase gene GmARI1 enhances aluminum tolerance in Arabidopsis.

Zhang X, Wang N, Chen P, Gao M, Liu J, Wang Y, Zhao T, Li Y, Gai J - PLoS ONE (2014)

Performance of the transgenic Arabidopsis under 15 µM Al treatment.A. Root phenotypes of the 35S: GmARI1 overexpression lines GmARI1-3 and the wild type Col-0 grown on 15 µM Al in 1/2 MS (+Al, pH4.3), and 0 µM Al in 1/2 MS (-Al, pH4.3). B. Relative root growth (RRG, %) of the transgenic Arabidopsis lines (GmARI1-1, GmARI1-2, GmARI1-3) and the wild type (Col-0). RRG was calculated by the root growth length under Al treatment (15 µM Al in 1/2 MS, pH4.3) divided by the root growth length under control (0 µM Al in 1/2 MS, pH4.3). Error bars represent the standard error (SE), ** indicate the significance level of 0.01 by t-tests.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4218711&req=5

pone-0111120-g009: Performance of the transgenic Arabidopsis under 15 µM Al treatment.A. Root phenotypes of the 35S: GmARI1 overexpression lines GmARI1-3 and the wild type Col-0 grown on 15 µM Al in 1/2 MS (+Al, pH4.3), and 0 µM Al in 1/2 MS (-Al, pH4.3). B. Relative root growth (RRG, %) of the transgenic Arabidopsis lines (GmARI1-1, GmARI1-2, GmARI1-3) and the wild type (Col-0). RRG was calculated by the root growth length under Al treatment (15 µM Al in 1/2 MS, pH4.3) divided by the root growth length under control (0 µM Al in 1/2 MS, pH4.3). Error bars represent the standard error (SE), ** indicate the significance level of 0.01 by t-tests.
Mentions: The gene expression of GmARI1 was induced early by Al stress, therefore we further studied its function in transgenic plants. An expression plasmid vector of pMDC83-GmARI1 was constructed and introduced into Arabidopsis plants using floral dip method. Transgenic T3 Arabidopsis over-expressing GmARI1 were generated and the positive transgenic lines were identified by RT-PCR (Fig. S2). Seeds of three T3 homozygous transgenic lines (GmARI1-1, GmARI1-2, and GmARI1-3) and wild type Col-0 were germinated on 1/2 MS medium. After 10 days, the seedlings were transferred to 1/2 MS (pH4.3) containing 15 µM Al or 0 µM Al as a control. Fifteen days later, the root growth of the wild type Col-0 was severely inhibited by 15 µM Al as compared with the control medium (0 µM), while the transgenic lines were little affected by Al (Fig. 9 A). The relative root growth (RRG) of the transgenic lines was significantly (p<0.01) longer than the wild type under Al treatment (Fig. 9B). The relative abundance of GmARI1 in the transgenic lines of GmARI1-1 and GmARI1-2 is higher than GmARI1-3, which coincided with RRG result (Fig. S3 and Fig. 9B).

Bottom Line: In vitro ubiquitination assay showed GmARI1 protein has E3 ligase activity.Overexpression of GmARI1 significantly enhanced the aluminum tolerance of transgenic Arabidopsis.These findings suggest that GmARI1 encodes a RBR type E3 ligase, which may play important roles in plant tolerance to aluminum stress.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Crop Genetics and Germplasm Enhancement, National Center for Soybean Improvement, Key Laboratory for Biology and Genetic Improvement of Soybean (General, Ministry of Agriculture), Nanjing Agricultural University, Nanjing, Jiangsu, China.

ABSTRACT
Ariadne (ARI) subfamily of RBR (Ring Between Ring fingers) proteins have been found as a group of putative E3 ubiquitin ligases containing RING (Really Interesting New Gene) finger domains in fruitfly, mouse, human and Arabidopsis. Recent studies showed several RING-type E3 ubiquitin ligases play important roles in plant response to abiotic stresses, but the function of ARI in plants is largely unknown. In this study, an ariadne-like E3 ubiquitin ligase gene was isolated from soybean, Glycine max (L.) Merr., and designated as GmARI1. It encodes a predicted protein of 586 amino acids with a RBR supra-domain. Subcellular localization studies using Arabidopsis protoplast cells indicated GmARI protein was located in nucleus. The expression of GmARI1 in soybean roots was induced as early as 2-4 h after simulated stress treatments such as aluminum, which coincided with the fact of aluminum toxicity firstly and mainly acting on plant roots. In vitro ubiquitination assay showed GmARI1 protein has E3 ligase activity. Overexpression of GmARI1 significantly enhanced the aluminum tolerance of transgenic Arabidopsis. These findings suggest that GmARI1 encodes a RBR type E3 ligase, which may play important roles in plant tolerance to aluminum stress.

Show MeSH
Related in: MedlinePlus