A novel aldo-keto reductase gene, <i>IbAKR,</i> from sweet potato confers higher tolerance to cadmium stress in tobacco

doi:10.15302/J-FASE-2018225

Front. Agr. Sci. Eng.

2018, Vol. 5

Issue (2) : 206-213 https://doi.org/10.15302/J-FASE-2018225

RESEARCH ARTICLE

A novel aldo-keto reductase gene, IbAKR, from sweet potato confers higher tolerance to cadmium stress in tobacco

Jinxi HUO, Bing DU, Sifan SUN, Shaozhen HE, Ning ZHAO, Qingchang LIU, Hong ZHAI(

)

Key Laboratory of Sweetpotato Biology and Biotechnology, Ministry of Agriculture/Beijing Key Laboratory of Crop Genetic Improvement/Laboratory of Crop Heterosis and Utilization, Ministry of Education, China Agricultural University, Beijing 100193, China

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Abstract

High concentrations of Cd can inhibit growth and reduce the activity of the photosynthetic apparatus in plants. In several plant species, aldo-keto reductases (AKRs) have been shown to enhance tolerance to various abiotic stresses by scavenging cytotoxic aldehydes; however, few AKRs have been reported to enhance Cd stress tolerance. In this study, the gene IbAKR was isolated from sweet potato. The relative expression levels of IbAKR increased significantly (approximately 3-fold) after exposure to 200 mmol·L⁻¹ CdCl₂ or 10 mmol·L⁻¹ H₂O₂. A subcellular localization assay showed that IbAKR is predominantly located in the nucleus and cytoplasm. IbAKR-overexpressing tobacco plants showed higher tolerance to Cd stress than wild-type (WT). Transgenic lines showed a significant ability to scavenge malondialdehyde (MDA) and methylglyoxal (MG). In addition, proline content and superoxide dismutase activity were significantly higher and H₂O₂ levels were significantly lower in the transgenic plants than in the WT. Quantitative real-time PCR analysis showed that the reactive oxygen species (ROS) scavenging genes encoding guaiacol peroxidase (GPX), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR) and peroxidase (POD) were significantly upregulated in transgenic plants compared to WT under Cd stress. These findings suggest that overexpressing IbAKR enhances tolerance to Cd stress via the scavenging of cytotoxic aldehydes and the activation of the ROS scavenging system.

Keywords cadmium stress IbAKR Ipomoea batatas sweet potato

Corresponding Author(s): Hong ZHAI

Just Accepted Date: 19 April 2018 Online First Date: 16 May 2018 Issue Date: 28 May 2018

Cite this article:

Jinxi HUO,Bing DU,Sifan SUN, et al. A novel aldo-keto reductase gene, IbAKR, from sweet potato confers higher tolerance to cadmium stress in tobacco[J]. Front. Agr. Sci. Eng. , 2018, 5(2): 206-213.

URL:

https://academic.hep.com.cn/fase/EN/10.15302/J-FASE-2018225
https://academic.hep.com.cn/fase/EN/Y2018/V5/I2/206

Fig.1 Analysis of IbAKR expression. (a) Organ-specific expression assay of IbAKR; (b,c) expression levels of IbAKR after treatment with 200 mmol·L⁻¹ CdCl₂(b) or 10 mmol·L⁻¹ H₂O₂(c). Data are presented as the mean±SE (n = 3). * and ** indicate a significant difference from 0 h at P<0.05 and<0.01, respectively, by Student’s t-test.

Fig.2 Subcellular localization of the IbAKR-GFP fusion protein in onion epidermal cells. The empty 35S:GFP vector (GFP) (as a control) and 35S:IbAKR-GFP (IbAKR-GFP) were transiently expressed in onion epidermal cells. Fluorescence from AKR-GFP and GFP were observed using a confocal laser scanning microscope under 488 nm excitation. Bar= 100 mm.

Fig.3 Elevated cadmium tolerance in IbAKR-OE tobacco plants in the in vitro assay. (a) In vitro-grown tobacco plants were transferred to 1/2 MS medium with or without 200 mmol·L⁻¹ CdCl₂. Photographs were taken 4 weeks after the start of treatment; (b, c) Fresh weight (b) and root length (c) were measured. Data are presented as the mean±SE (n = 3). * and ** indicate a significant difference from WT at P<0.05 and<0.01, respectively, by Student’s t-test.

Fig.4 Elevated cadmium tolerance in IbAKR-OE tobacco plants grown in pots. (a) Eight-week-old plants in a greenhouse were irrigated every 3 d with Hoagland solution containing 100 mmol·L⁻¹ CdCl₂ and morphological changes in the three lines (WT, L1, L2 from left to right) were photographed before and 2 weeks after the start of treatment; red circles indicate necrotic spots; (b–e) content of stress-related metabolites and superoxide dismutase (SOD) in transgenic lines and WT after treatment with 100 mmol·L⁻¹ CdCl₂ for 2 weeks. Data are presented as the mean±SE (n = 3). * and ** indicate a significant difference from WT at P<0.05 and<0.01, respectively, by Student’s t-test.

Fig.5 Leaf disk senescence under methylglyoxal (MG) stress as measured by chlorophyll content. Chlorophyll content (mg·g⁻¹FW) of leaf discs from WT and transgenic line L2 tobacco plants, kept for 3 d at different concentrations of MG. Data are presented as the mean±SE (n = 3). * and ** indicate a significant difference from WT at P<0.05 and<0.01, respectively, by Student’s t-test.

Fig.6 Analysis of the expression of ROS scavenging-related genes in transgenic plants and WT. The relative expression levels of genes encoding guaiacol peroxidase (GPX), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR) and peroxidase (POD) were analyzed after treatment with 100 mmol·L⁻¹ CdCl₂ for 2 weeks. Data are presented as the mean±SE (n = 3). * and ** indicate a significant difference from WT at P<0.05 and<0.01, respectively, by Student’s t-test.

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