Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX 79409, USA.
Plant Cell Physiol. 2011 Jan;52(1):149-61. doi: 10.1093/pcp/pcq182. Epub 2010 Nov 18.
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.
通过 sos3（盐过度敏感 3）抑制子筛选，回收了两个等位基因抑制子突变体，它们是强 sos3 抑制子 sos3hkt1-1 的弱等位基因。分子特征鉴定表明，这两个弱 sos3 抑制子中的 T-DNA 插入到拟南芥 HKT1（AtHKT1，At4g10310）的远端启动子区域，导致串联重复与 AtHKT1 启动子的近端区域物理分离。串联重复位于 ATG 起始密码子上游约 3.9kb，作为增强子元件促进报告基因表达。ATG 起始密码子上游约 2.6kb 的假定小 RNA 靶区高度甲基化。在小 RNA 生物发生突变体 rdr2 中，CHG 和 CHH 甲基化而非 CG 甲基化显著减少，表明该区域的非 CG 甲基化是由小 RNA 介导的。在 rdr2 中分析 AtHKT1 的表达表明，假定的小 RNA 靶区中的非 CG 甲基化抑制了地上部分的 AtHKT1 表达。DNA 甲基化缺陷突变体 met1-3 在假定的小 RNA 靶区中几乎完全丧失了总胞嘧啶甲基化，对盐胁迫敏感。假定的小 RNA 靶区和串联重复对于维持 AtHKT1 表达模式至关重要，这些表达模式对于盐耐受性至关重要。
