Promoter Cis-Element Analyses Reveal the Function of αVPE in Drought Stress Response of Arabidopsis

Abstract

Vacuolar processing enzyme (VPE) is a cysteine protease responsible for vacuolar proteins' maturation and regulation of programmed cell death (PCD). Four isoforms of Arabidopsis thaliana VPEs were identified previously, but only the functions of βVPE, γVPE, and δVPE were determined. The specific function of a gene is linked to the cis-acting elements in the promoter region. A promoter analysis found repetitive drought-related cis-elements in αVPE, which highlight its potential involvement in drought regulation in A. thaliana. The further co-expression network portraying genes interacting with αVPE substantiated its drought-regulation-related function. Expression of αVPE was upregulated after drought treatment in A. thaliana. To confirm the role of αVPE, a loss of function study revealed that αVPE knockout mutants remained green compared with WT after drought treatment. The mutants had reduced proline activity, decreased sucrose content, and lower MDA content, but increased photosynthetic pigments, indicating that αVPE negatively regulates drought tolerance in A. thaliana. Taken together, our findings serve as important evidence of the involvement of αVPE in modulating drought tolerance in A. thaliana.

Keywords: drought; motifs; stress; vacuolar processing enzyme.

Figure 1

Predicted cis-element analysis in the promoter region of αVPE and co-expression networks for αVPE assembled from transcriptome data in ATTED-II with default parameters. (a) Promoter sequences (2.5 kb) of αVPE were analyzed by PlantCARE and PLACE Web Signal Scan. Different colour and shape boxes stand for different cis-elements. Asterisk symbol represents the cis-element in an inverted direction on the promoter region. (b) Bolder lines show a direct connection with αVPE, while dotted lines show an indirect connection of co-expressed genes with αVPE.

Figure 2

Normalised relative gene expression of the AtVPE family (αVPE, βVPE, γVPE, and δVPE) in WT upon drought treatment. Data indicate the mean (±SD) of three biological replicates. Asterisk symbol represents significant difference at p < 0.05 compared with the control.

Figure 3

Effects on WT and alpha mutant upon drought treatment. Morphology of (a) WT before drought treatment and (b) alpha mutants before drought treatment. (c) WT after drought treatment. (d) Alpha mutants after drought treatment. (e) RWC (%) in leaves of both WT and alpha mutants after drought treatment. (f) Total proline activity was measured in nM/gFW. (g) Total sugar content was measured in mg/gFW. (h) Total MDA content was measured in nM/gFW. (i) Total photosynthetic pigment content was measured in mg/gFW. The results indicate the mean (±SD) of three biological replicates. Asterisk symbol represents a significant difference at p < 0.05 compared with wild type. Double asterisk symbols represent significant difference at p < 0.01 compared with WT.