Comparative and functional promoter analysis of desiccation-related genes from three closely related plant species differing in desiccation tolerance

Abstract

The closely related resurrection plants C. plantagineum and L. brevidens have the rare ability to withstand desiccation of their vegetative tissues. The species L. subracemosa is a close relative of C. plantagineum and L. brevidens, but cannot tolerate desiccation of its vegetative tissues. This variability in desiccation tolerance was used to assess differences in the transcriptional regulation of desiccation-responsive genes between C. plantagineum, L. brevidens and L. subracemosa. Comparative analysis of the dehydration-responsive transcriptomes of L. brevidens and L. subracemosa revealed that L. brevidens expressed more transcripts associated with desiccation tolerance. This finding indicates that the expression of dehydration-responsive genes is regulated differentially between the desiccation tolerant plant L. brevidens and the desiccation sensitive plant L. subracemosa. Transcript encoding LEA proteins were highly abundant in the dehydration-responsive transcriptome of L. brevidens. <br /> A promoter study was conducted to gain more understanding of the transcriptional regulation of dehydration-responsive genes in C. plantagineum, L. brevidens and L. subracemosa. Promoter regions of five dehydration-responsive genes were analyzed: LEA 6-19, LEA-like 11-24, LEA 3-06, LEA 27-45 and DSP22, respectively. Putative cis-acting regulatory elements known to mediate ABA- and dehydration-responses were detected in promoter regions of all genes. Comparison of promoter regions revealed that several of the detected cis-acting regulatory elements were conserved among the promoters of C. plantagineum, L. brevidens and L. subracemosa. Expression analysis showed that the LEA-like 11-24 gene was differentially expressed in C. plantagineum, L. brevidens and L. subracemosa in response to different stress treatments. The activity of LEA-like 11-24 promoter fragments of C. plantagineum, L. brevidens and L. subracemosa was analyzed in a transient expression assay. Functional promoter analysis demonstrated differences in promoter activity between LEA-like 11-24 promoter fragments of C. plantagineum, L. brevidens and L. subracemosa. It was shown that variation in promoter architecture accounts for differences in LEA-like 11-24 promoter activity between C. plantagineum, L. brevidens and L. subracemosa. <br /> A mutagenesis approach was employed to identify cis-acting regulatory elements essential for promoter activity. Cis-acting regulatory elements critical for ABA- and dehydration-responsive promoter activity were detected in LEA-like 11-24 promoter fragments of C. plantagineum and L. brevidens. Furthermore, it was demonstrated that specific cis-acting regulatory elements were structurally and functionally conserved among the LEA-like 11-24 promoters of C. plantagineum and L. brevidens. <br /> The yeast one hybrid system showed that the bZIP1 protein of C. plantagineum was able to interact with the C. plantagineum LEA-like 11-24 promoter. This finding indicates that the C. plantagineum bZIP1 protein is involved in transcriptional regulation of the LEA-like 11-24 gene. It was shown that the C. plantagineum bZIP1 protein was nuclear localized in epidermal leaf cells of C. plantagineum. Expression analysis revealed that the bZIP1 gene was constitutively expressed in C. plantagineum leaf tissue. Although co-expression of bZIP1 did not enhance the promoter activity of the LEA-like 11-24 promoter fragment in C. plantagineum leaves, it was proposed that the bZIP1 protein is involved, but not sufficient for the activation of LEA-like 11-24 gene expression in C. plantagineum. This study provided more insight in the regulation of desiccation-responsive genes in the closely related species C. plantagineum, L. brevidens and L. subracemosa.