Isolation and characterization of maize ZmPP2C26 gene promoter in drought-response

Abstract

The clade A members of serine/threonine protein phosphatase 2Cs (PP2Cs) play crucial roles in plant growth, development, and stress response via the ABA signaling pathway. But little is known about other PP2C clades in plants. Our previous study showed that maize the ZmPP2C26, a clade B member of ZmPP2Cs, negatively regulated drought tolerance in transgenic Arabidopsis. However, the upstream regulatory mechanism of ZmPP2C26 remains unclear. In the present study, the expression of ZmPP2C26 gene in maize was analyzed by quantitative real time PCR (qRT-PCR). The results showed that the expression of ZmPP2C26 in shoot and root was both significantly inhibited by drought stress. Subsequently, a 2175 bp promoter of ZmPP2C26 was isolated from maize genome (P ₂₁₇₅). To validate whether the promoter possess some key cis-element and negatively drive ZmPP2C26 expression in drought stress, three 5´-deletion fragments of 1505, 1084 and 215 bp was amplified from P ₂₁₇₅ and were fused to β-glucuronidase (GUS) and luciferase gene (LUC) to produce promoter::GUS and promoter::LUC constructs, and transformed into tobacco, respectively. Transient expression assays indicated that all promoters could drive GUS and LUC expression. The GUS and LUC activity were both significantly inhibited by PEG-6000 treatment. Notably, the - 1084 to - 215 bp promoter possess one MBS element and inhibits the expression of GUS and LUC under drought stress. Meanwhile, we found that the 215 bp length is enough to drive ZmPP2C26 expression. These findings will provide insights into understanding the transcription-regulatory mechanism of ZmPP2C26 negatively regulating drought tolerance.

Keywords: Drought stress; Maize; Promoter; Serine/threonine protein phosphatase 2C.