Regulated AtHKT1 Gene Expression by a Distal Enhancer Element and DNA Methylation in the Promoter Plays an Important Role in Salt Tolerance

Abstract

Through sos3 (salt overly sensitive 3) suppressor screening, two allelic suppressor mutants that are weak alleles of the strong sos3 suppressor sos3hkt1-1 were recovered. Molecular characterization identified T-DNA insertions in the distal promoter region of the Arabidopsis thaliana HKT1 (AtHKT1, At4g10310) in these two weak sos3 suppressors, which results in physical separation of a tandem repeat from the proximal region of the AtHKT1 promoter. The tandem repeat is approximately 3.9 kb upstream of the ATG start codon and functions as an enhancer element to promote reporter gene expression. A putative small RNA target region about 2.6 kb upstream of the ATG start codon is heavily methylated. CHG and CHH methylation but not CG methylation is significantly reduced in the small RNA biogenesis mutant rdr2, indicating that non-CG methylation in this region is mediated by small RNAs. Analysis of AtHKT1 expression in rdr2 suggests that non-CG methylation in the putative small RNA target region represses AtHKT1 expression in shoots. The DNA methylation-deficient mutant met1-3 has nearly complete loss of total cytosine methylation in the putative small RNA target region and is hypersensitive to salt stress. The putative small RNA target region and the tandem repeat are essential for maintaining AtHKT1 expression patterns crucial for salt tolerance.

Salt Stress Induces Non-CG Methylation in Coding Regions of Barley Seedlings (Hordeum vulgare)

“…This is in accordance with our finding that dose-insensitive salt-induced DMMs appeared more commonly in sites that could facilitate perturbation in the expression of genes that hypothetically could be part of a molecular response to salinity, as reported elsewhere [65]. There is evidence from previous studies to suggest that salt-induced DMMs can play important roles in evoking metabolic differences between seedlings growing under control and saline conditions [25,26,29,34,64,66], although the markers found in these works were either few in number or else identified from a single genotype. The provision here of a robust list of consistent, salt-responsive DMMs therefore provides a useful starting point from which to gather a more holistic picture of DNA methylation-mediated regulation of molecular responses to salt exposure.…”

“…Given that salt-induced DMMs have the potential to regulate gene expression, the functions of differentially methylated genes were explored for possible correlations with stress responsive genes. The correlation of DM genes with GO terms that are related to plant responses to stress, such as "negative regulation of signal transduction", "photosynthesis", "response to osmotic stress" and "ion transmembrane transport", is at least consistent with the possibility that salt-induced DMMs targeted genes which could play active functions in a plant's response to salt, in broad accordance with previously expression studies of salt response [29,[81][82][83]. In addition, some of the DM genes identified here were enriched for GO terms, such as "hydrolase activity", "oxidoreductase activity", "nucleic acid binding", and "translation factor activity".…”

HKT transporters mediate salt stress resistance in plants: from structure and function to the field

“…The AtHKT1;1 promoter sequence contains a small RNA binding region À2.6 kb upstream of the AtHKT1;1 start codon, which was found to be more strongly methylated in leaves compared to roots (Figure 3a). Differences in methylation may contribute to higher level of AtHKT1;1 expression in roots [48]. As another transcriptional cis regulatory element in the AtHKT1;1 promoter, a direct repeat sequence located À5.3 kb to À3.9 kb upstream of the transcriptional start codon of AtHKT1;1 was found through the comparative approach on salt sensitivity in several Arabidopsis cultivars ( Figure 3) [41,48].…”

Flexible shift on gene body methylation and transcription of LpCYP72A161 exposed to temperature stress in perennial ryegrass

“…Abiotic stress's impact on DNA methylation has been widely reported in plants at the genome level (Ci et al, 2015;Gayacharan and Joel 2013;Tang et al, 2014b;Wang et al, 2015a), and on the promoter regions (Baek et al, 2011;Chinnusamy and Zhu 2009), but only a few studies focused on changes in DNA methylation on gene body regions. Currently, the relationship between gene body methylation and transcription remains unclear (Maunakea et al, 2010;Zemach et al, 2010).…”