Identification Of CPA Gene Family In Phyllostachys Edulis And The Functional Analysis Of PheNHX2

Moso bamboo(Phyllostachys edulis)is one of the most important non-wood forest products in the world,which has high cultural,ecological and economic values.While,many adverse environmental factors such as low temperature,drought and salinity seriously restrict the development of bamboo industry.Plant monovalent cation/proton antiporters(CPAs)are types of ion transport-related transmembrane transporters,which play an important role in plant salt stress response.Plant CPAs include three types:Na~+/H~+antiporter(NHX),K~+efflux antiporter(KEA)and cation/H~+exchanger(CHX).At present,the CPA genes have been isolated and identified in many other species,while the researches about the molecular characteristics of CPA genes in moso bamboo are few.In this study,the CPA genes of moso bamboo were identified at the whole genome level through using bioinformatics methods.Then,the tissue and induced expression patterns of moso bamboo NHX-type genes were analyzed by qRT-PCR,and the stress-resistance candidate gene PheNHX2 was screened out.The Agrobacterium-mediated method was further used to overexpress PheNHX2 in Arabidopsis thaliana to study the biological function of PheNHX2.The main results were obtained:1.37 CPA genes containing the Na＿H＿Exchanger(PF00999)domain were identified from the moso bamboo genome.Phylogenetic analysis showed that the CPA genes of moso bamboo can be divided into three categories:10 NHX genes(PheNHX1-PheNHX10),6 KEA genes(PheKEA1-PheKEA6)and 21 CHX genes(PheCHX1-PheCHX21).The analysis of protein conserved domains predicted 10conserved motifs in moso bamboo CPA proteins.Among them,the Motif 2 belonging to the fragment of the Na＿H＿Exchanger domain was highly conserved and distributed in all CPA members.2.The tissue expression pattern of moso bamboo NHX genes showed that the NHX genes were specifically expressed in multiple tissues.And the induced expression pattern showed that salt treatment can induce the expression of NHX genes in different degrees.Among them,the expression of PheNHX2 was significantly up-regulated under NaCl treatment.3.The subcellular localization experiment of PheNHX2 gene showed that PheNHX2 was located on the vacuolar membrane.Overexpression PheNHX2 transgenic Arabidopsis under salt stress significantly inhibits plant seed germination and root growth,affects the balance of Na~+and K~+in the cell,and reduces the salt tolerance of transgenic Arabidopsis.At the same time,the measures of the chlorophyll content,malondialdehyde content,peroxidase and catalase activity,relative conductivity and other related physiological indicators further confirmed that overexpression of PheNHX2 reduced the salt tolerance of transgenic Arabidopsis.The qRT-PCR experiments showed that under salt stress,overexpression of PheNHX2 significantly inhibited the expression of some genes related to stress and ion transport in transgenic Arabidopsis.Moso bamboo NHX genes are types of genes related to plant salt stress response.PheNHX2,which located on the tonoplast membrane,negatively regulates the salt tolerance of plants.The results laid the foundation for the research on the function of the CPA gene of moso bamboo and the mechanism of PheNHX2 regulating salt tolerance.