COX, Harvey, Turning East : The Promise and Peril of the New Orientalism

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

Expression profiles of <i>TPS4</i> in rapeseed subjected to salinity stress.

<p>Salinity-stressed shoots (A and B) and roots (C and D) of two rapeseed cultivars, Exagone (salt-tolerant) and Toccata (salt-sensitive), were used to quantify levels of <i>TPS</i>4 transcripts relative to those in unstressed control plants. Real-time PCR analysis was performed using gene-specific primers. The expression of the <i>TPS</i>4 gene was normalised relative to the expression of internal control genes, which encode glyceraldehyde-3-phosphate dehydrogenase (<i>GAPDH</i>) and elongation factor 1-alpha (<i>EF-1</i>-α. Error bars indicate the standard error calculated using a 95% confidence interval. Relative expression values (fold change) are on a logarithm base 2 scale.</p

Stages of collection of seedlings.

<p>Seeds were sterilised in 0.1% NaClO, incubated in distilled water (0 mmol/L), and placed on moistened filter paper. Four days after sowing (4 DAS), seedlings (A) were split and grown in boxes containing distilled water (control; B, D, F, H, J, L, O) or NaCl solution (100 mmol/L; C, E, G, I, K). Moreover, at 14 DAS samples grown under stressed conditions (K) were split: half were kept under stressed conditions (N and Q) and the other half were grown under control conditions (recovery; M and P). All samples were collected at 8 am, except for samples indicated with the letter e (evening) which were collected at 8 pm. Samples 4e, 5 and 5e were used for qRT-PCR and not for MSAP analysis.</p

Expression profiles of <i>Lacerata</i> (<i>LCR</i>) in rapeseed after exposure to salinity stress.

<p>Salinity-stressed shoots (A and B) and roots (C and D) of two rapeseed cultivars, Exagone (salt-tolerant, A and C) and Toccata (salt-sensitive, B and D), were used to quantify the expression of <i>LCR</i> relative to control. Real-time PCR analysis was performed using gene-specific primers. The expression of <i>LCR</i> was normalised relative to the expression of internal control genes, which encode glyceraldehyde-3-phosphate dehydrogenase (<i>GAPDH</i>) and elongation factor 1-alpha (<i>EF-1</i>- α. Error bars indicate the standard error calculated using a 95% confidence interval. Relative expression values (fold change) are on a logarithm base 2 scale.</p