Heterologous expression of the cysteine proteinase gene VaCP17 from the cold-adapted grapevine Vitis amurensis increases cold tolerance in Arabidopsis

Xin Shu,Bao Gu,Yunyun Tang,Hongjuan Zhang,Jianxia Zhang

doi:10.1007/s11240-022-02256-x

Abstract

Cysteine proteinases (thiol) carry out diverse and critical functions in plants through their ability to hydrolyze peptide bonds in target proteins. Here, we cloned a cysteine proteinase gene designated VaCP17 from a highly cold-resistant wild Vitis amurensis accession ‘Shuangyou’, and then its potential function in cold resistance was investigated. The results showed that the CDS of VaCP17 is 1404 bp, encoding 467 amino acids, the VaCP17 protein localized to the cell membrane. Expression of CP17 was highly distinctive among different structures of ‘Shuangyou’ and the cold-sensitive Vitis vinifera cultivar ‘Red Globe’, with the highest expression in the stem of ‘Shuangyou’ and the flower of ‘Red Globe’. Arabidopsis plants constitutively expressing a VaCP17-GREEN FLUORESCENT PROTEIN fusion (35S::VaCP17-GFP) showed increased survival after transient exposure to freezing (-6 °C), and showed lower electrolyte leakage and MDA content, higher soluble sugar content and SOD, POD and CAT activities, as compared with non-transgenic Arabidopsis controls. The expression of nine cold-resistance related genes (CBF1, CBF2, CBF3, RD29A, COR15A, KIN1, NCED3, AOC1 and JAZ10) in 35S::VaCP17-GFP plants was increased under cold treatment at 4 °C, relative to control plants. Using a yeast two-hybrid system, we identified VaNAC72, VaCAM7 and VaDi19 as potential interactors of VaCP17, and their interactions were demonstrated by a bimolecular fluorescence complementation assay. In conclusion, we revealed that VaCP17 can enhance cold resistance by influencing physiology and biochemistry and the expression of cold resistance related genes under cold stress.

Highlights

Cold is one of the most important abiotic stresses limiting plant geographical distribution and crop production (Ye et al 2017)
The results showed that the coding sequence (CDS) of VaCP17 is 1404 bp, encoding 467 amino acids, the VaCP17 protein localized to the cell membrane
Cysteine proteinases play important roles in plant growth and development, senescence and programmed cell death, and it has been reported that they may participate in response to various stresses (Xu et al 2003; Zang et al 2010; Zheng et al 2018)

Summary

Introduction

Cold is one of the most important abiotic stresses limiting plant geographical distribution and crop production (Ye et al 2017). Jasmonic acid induces the degradation of JAZ protein and activates the expression of cold resistance-related genes, improving plant cold resistance (An et al 2021). Abiotic stresses such as cold, salt and drought usually increase the content of ABA, which in turn induces stomatal closure and reduces transpiration rate; ABA can regulate the expression of stress-related genes (Mulholland et al 2003; Jiang et al 2014; Li et al 2019). The rate-limiting enzyme of ABA biosynthesis is 9-cisepoxycarotenoid dioxygenases (NCEDs) Together, these mechanisms act to enhance the stability of cellular membranes by promoting accumulation of osmolytes such as proline, soluble sugar and soluble protein (Ashraf and Foolad 2007)

Methods

Results

Conclusion