The transcriptional regulatory mechanism of the peroxisomal ascorbate peroxidase (pAPX) gene cloned from an extreme halophyte, Salicornia brachiata.

摘要

Peroxisomal ascorbate peroxidase detoxifies H2O2 leaching out from peroxisomes into the cytoplasm. The present study describes transcript expression and cis-regulation of the SbpAPX gene cloned from an extreme halophyte, Salicornia brachiata, in the steady state and under different stresses. About 2-fold elevated transcript expression was found in salt- and drought-treated shoots at 12 h compared with control, while 1.9-fold increased expression was observed under heat treatment. In roots, the transcript level was down-regulated at 2 h, thereafter increasing with the time of exposure and reaching a maximum at the control level. The SbpAPX promoter has characteristic cis-regulatory ABA-dependent abiotic stress-responsive elements. The full-length promoter (1,024 bp, PP1) and deletion constructs -838 (PP2), -697 (PP3), -433 (PP4) and -185 bp (PP5) were fused with the GUS ( beta -glucuronidase) gene and transformed into tobacco for functional validation. Expression of GUS increased significantly in transgenic plants under stress. Quantitative expression analysis of GUS in T1 plants revealed that promoter PP5 is efficient for gene expression. In planta transient expression further suggested that the promoter PP5 contains efficient stress-inducible elements. A steep decline in GUS expression in PP3, and thereafter an elevated expression in PP4 and PP5, suggested the presence of a repressor element between -696 and -433 bp, while an enhancer element was predicted between -838 and -697 bp. Further, transient expression analyses and electrophoretic mobility shift assay revealed that the core sequence of cis-acting motifs ATAA and CCTCAA function as enhancer and repressor binding sites, respectively. Based on the study, a model is proposed for the cis-regulation of the SbpAPX gene. The present study provides a useful insight for understanding gene expression regulation in a halophyte with or without stress. Furthermore, potential stress-responsive promoter-driven expression of introgressed gene(s) can be used for engineering crops with enhanced stress tolerance.