Characterization of a Stress-Enhanced Promoter from the Grass Halophyte, Spartina alterniflora L.

SIMPLE SUMMARY: Promoter is an important component to drive the expression of desirable genes in the genetic modification of plants. Constitutive promoters that direct the expression of genes at all times in all tissues may affect the growth and development of plants. Therefore, there is a need to increase the availability of promoters that are induced by different environmental stresses and/or developmental stages. Extremophiles, such as halophytes, which thrive under harsh environmental stresses, are considered important sources of such stress-inducible (or stress-enhanced) promoters. To this end, we isolated the promoter region of a stress-responsive gene SaAsr1 from a grass halophyte smooth cordgrass. Characterization of the promoter and its deletion derivatives identified several regulatory elements that contribute to the expression of a gene at varying degrees under salt and drought stress. A known stress-responsive gene expressed under the control of the stress-enhanced promoter reported here produced plants that were normal and healthy in comparison with constitutive promoters that produced plants with compromised growth. Our results further validated the reports that gene expression under stress-inducible/enhanced promoter is a better strategy for genetic engineering to develop stress-resilient crop plants. ABSTRACT: Stress-inducible promoters are vital for the desirable expression of genes, especially transcription factors, which could otherwise compromise growth and development when constitutively overexpressed in plants. Here, we report on the characterization of the promoter region of a stress-responsive gene SaAsr1 from monocot halophyte cordgrass (Spartina alterniflora). Several cis-acting elements, such as ABRE (ABA-responsive element), DRE-CRT (dehydration responsive-element/C-Repeat), LTRE (low temperature-responsive element), ERE (ethylene-responsive element), LRE (light-responsive element), etc. contributed at varying degrees to salt-, drought- and ABA-enhanced expression of gusA reporter gene in Arabidopsis thaliana under the full-length promoter, pAsr1(1875) and its deletion derivatives with an assortment of cis-regulatory motifs. The smallest promoter, pAsr1(491), with three cis-acting elements (a CCAAT box-heat responsive, an LRE, and a copper responsive element) conferred drought-enhanced expression of gusA; pAsr1(755) (with an ABRE and a DRE) presented the highest expression in ABA and drought; and pAsr1(994) with seven ABREs and two DREs conferred optimal induction of gusA, especially under drought and ABA. Arabidopsis transgenics expressing a known abiotic stress-responsive gene, SaADF2 (actin depolymerization factor 2), under both pAsr1(1875) and p35S promoters outperformed the wild type (WT) with enhanced drought and salt tolerance contributed by higher relative water content and membrane stability with no significant difference between pAsr1(1875):SaADF2 or p35S:SaADF2 lines. However, pAsr1(1875):SaADF2 lines produced healthy plants with robust shoot systems under salt stress and control compared to slightly stunted growth of the p35S:SaADF2 plants. This reestablished the evidence that transgene expression under a stress-inducible promoter is a better strategy for the genetic manipulation of crops.