Abstract
Ubiquitination is a common regulatory mechanism, playing a critical role in diverse cellular and developmental processes in eukaryotes. However, a few reports on the functional correlation between E3 ubiquitin ligases and reactive oxygen species (ROS) or reactive nitrogen species (RNS) metabolism in response to stress are currently available in plants. In the present study, the E3 ubiquitin ligase gene AdBiL (Adi3 Binding E3 Ligase) was introduced into tomato line Ailsa Craig via Agrobacterium-mediated method. Transgenic lines were confirmed for integration into the tomato genome using PCR. Transcription of AdBiL in various transgenic lines was determined using real-time PCR. Evaluation of stress tolerance showed that T1 generation of transgenic tomato lines showed only mild symptoms of chilling injury as evident by higher biomass accumulation and chlorophyll content than those of non-transformed plants. Compared with wild-type plants, the contents of AsA, AsA/DHA, GSH and the activity of GaILDH, γ-GCS and GSNOR were increased, while H2O2, , MDA, NO, SNOs, and GSNO accumulations were significantly decreased in AdBiL overexpressing plants in response to chilling stress. Furthermore, transgenic tomato plants overexpressing AdBiL showed higher activities of enzymes such as G6PDH, 6PGDH, NADP-ICDH, and NADP-ME involved in pentose phosphate pathway (PPP). The transgenic tomato plants also exhibited an enhanced tolerance against the necrotrophic fungus Cladosporium fulvum. Tyrosine nitration protein was activated in the plants infected with leaf mold disease, while the inhibition could be recovered in AdBiL gene overexpressing lines. Taken together, our results revealed a possible physiological role of AdBiL in the activation of the key enzymes of AsA–GSH cycle, PPP and down-regulation of GSNO reductase, thereby reducing oxidative and nitrosative stress in plants. This study demonstrates an optimized transgenic strategy using AdBiL gene for crop improvement against biotic and abiotic stress factors.
Highlights
Ubiquitination is a common regulatory mechanism that occurs in all eukaryotes to regulate diverse cellular and developmental processes at post-translational level (Liu et al, 2013)
An expected 751 bp fragment was observed with the AdBiL gene specific primers, whereas no corresponding band was detected in the untransformed plants (Figure 2A)
Despite a keen interest in plant RING motif-harboring ubiquitin E3 ligases for their involvement in diverse cellular and developmental processes, a few reports on the functional correlation between RING E3 ligases and reactive oxygen species (ROS) or NO signal-mediated stress responses are currently available
Summary
Ubiquitination is a common regulatory mechanism that occurs in all eukaryotes to regulate diverse cellular and developmental processes at post-translational level (Liu et al, 2013). Number of genes encoding E3s is more than 1,200 in Arabidopsis Such a high number of genes relative to the other eukaryotes underline the importance of E3s in regulating plant processes. Many studies have demonstrated that E3s are key regulators in the response of plants to abiotic stress, hormone signaling, photomorphogenesis, cell cycling and plant–microbe interactions (Hervé et al, 2011; Ling and Jarvis, 2015). AtCHIP, a U-box-containing E3 ubiquitin ligase, plays a critical role in temperature stress tolerance in Arabidopsis. Overexpression of AtCHIP in Arabidopsis renders plants more sensitive to both low- and high-temperature stresses. Arabidopsis plants lacking ubiquitin E3 ligase Suppressor of Ppi Locus (SP1) are hypersensitive to salt, osmotic, and oxidative stresses, whereas plants overexpressing SP1 are considerably more stress tolerant than wild-type (Ling and Jarvis, 2015). AtATL80-overexpressors are significantly more sensitive to cold stress than wild-type plants, while the AtATL80 mutant lines exhibit an increased tolerance to cold stress (Suh and Kim, 2015)
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