Cloning and expression analysis of Aerenchyma development related ShRBOH1 gene in Saussurea inversa Raab-Straube

Abstract

Saussurea inversa Raab-Straube is a typical alpine plant with well-developed aerenchyma, which is its most representative structural feature. Studying the formation and development of aerenchyma in S. inversa is of great theoretical significance for understanding the adaptive mechanisms of alpine plants to the environment. In this study, the aerenchyma development-related gene ShRBOH1 (GenBank accession number OP727799) was cloned from S. inversa by RT-PCR with RACE technology. Sequence analysis revealed that the cDNA of ShRBOH1 was 2158 bp long including an open reading frame of 2031 bp that encodes 676 amino acids. Subcellular localization prediction indicated that the ShRBOH1 protein is located in the cytoplasmic membrane, consistent with the localization of the NADPH protein. A comparison of the amino acid sequence homology showed that ShRBOH1 had higher similarity to CcRBOH1 of Cynara cardunculus L. The qRT-PCR analysis demonstrated that the ShRBOH1 gene was expressed in roots, stems, and leaves with significantly lower expression level in stems compared to other tissues. The expression of the ShRBOH1 gene was down-regulated under UV stress, up-regulated under cold stress, and initially up-regulated, then down-regulated, and finally up-regulated again under hypoxia stress. By comparing the changes in reactive oxygen species (ROS) content under the three types of stresses, we found that significant changes only occurred under hypoxic stress and were positively correlated with the expression of ShRBOH1. We hypothesize that ShRBOH1 positively regulates the formation of aerenchyma through ROS mediation, suggesting that the aerenchyma is primarily an important structure for adapting to hypoxia stress in alpine environments.