Abstract
Excessive soil salinity is a major ecological and agronomical problem, the adverse effects of which are becoming a serious issue in regions where saline water is used for irrigation. Plants can employ regulatory strategies, such as DNA methylation, to enable relatively rapid adaptation to new conditions. In this regard, cytosine methylation might play an integral role in the regulation of gene expression at both the transcriptional and post-transcriptional levels. Rapeseed, which is the most important oilseed crop in Europe, is classified as being tolerant of salinity, although cultivars can vary substantially in their levels of tolerance. In this study, the Methylation Sensitive Amplified Polymorphism (MSAP) approach was used to assess the extent of cytosine methylation under salinity stress in salinity-tolerant (Exagone) and salinity-sensitive (Toccata) rapeseed cultivars. Our data show that salinity affected the level of DNA methylation. In particular methylation decreased in Exagone and increased in Toccata. Nineteen DNA fragments showing polymorphisms related to differences in methylation were sequenced. In particular, two of these were highly similar to genes involved in stress responses (Lacerata and trehalose-6-phosphatase synthase S4) and were chosen to further characterization. Bisulfite sequencing and quantitative RT-PCR analysis of selected MSAP loci showed that cytosine methylation changes under salinity as well as gene expression varied. In particular, our data show that salinity stress influences the expression of the two stress-related genes. Moreover, we quantified the level of trehalose in Exagone shoots and found that it was correlated to TPS4 expression and, therefore, to DNA methylation. In conclusion, we found that salinity could induce genome-wide changes in DNA methylation status, and that these changes, when averaged across different genotypes and developmental stages, accounted for 16.8% of the total site-specific methylation differences in the rapeseed genome, as detected by MSAP analysis.
Highlights
Excessive salt accumulation in soil is a major ecological and agronomical problem [1], the adverse effects of which are becoming a serious issue in regions where saline water is used for irrigation [2]
The extent of DNA methylation ranged from 45.85% (7 days after sowing (DAS)) to 47.67% (4 DAS) in Exagone and from 39.71% (14 DAS) to 43.55% (4 DAS) in Toccata samples (Table 1)
Salinity stress decreased the percentage of total methylated bands in tolerant Exagone, but increased it in sensitive Toccata, when levels of DNA methylation were compared with those in the respective unstressed control. These findings indicated opposite effects of salinity stress on DNA methylation in genotypes of B. napus with different levels of tolerance to salinity stress (Table S2)
Summary
Excessive salt accumulation in soil is a major ecological and agronomical problem [1], the adverse effects of which are becoming a serious issue in regions where saline water is used for irrigation [2]. More than 45 million hectares of irrigated land have been damaged by salt, and 1.5 million hectares are taken out of production each year as a result of high levels of soil salinity [3,4]. During the onset and development of salt stress, major processes such as photosynthesis, protein synthesis, and energy and lipid metabolism, are affected [5]. Ionic stress causes symptoms of toxicity, such as chlorosis and necrosis, due to high Na+ levels, which affects plants by disrupting protein synthesis and interfering with enzyme activity [7,8,9]. A high concentration of Na+ reduces growth and inhibits cell division and expansion [10]
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