# Genome-Wide Identification and Expression Analysis of NHX Gene Family in Ziziphus jujuba var. spinosa Under Salt and Drought Stress

## TL;DR

This study identifies and analyzes NHX genes in sour jujube, revealing their role in salt stress tolerance and providing insights for future research.

## Contribution

The first genome-wide analysis of the NHX gene family in Ziziphus jujuba var. spinosa under abiotic stress.

## Key findings

- Eight NHX genes were identified in sour jujube, distributed across six chromosomes and classified into three subfamilies.
- Most ZjNHX genes are up-regulated by salt stress, with ZjNHX1 showing a 40-fold increase in root expression within 3 hours.
- Promoter regions of ZjNHX genes contain cis-regulatory elements linked to stress adaptation and phytohormone signaling.

## Abstract

Background/Objectives: Ziziphus jujuba var. spinosa (sour jujube) is a traditional medicinal plant with remarkable tolerance to abiotic stresses, particularly salinity. However, the regulatory mechanisms underlying its salt stress tolerance remain unclear. NHX genes play a crucial role in plant adaptation to salt stress by mediating Na+/K+ transport to maintain intracellular ion homeostasis and pH balance. Although the NHX gene family has been characterized in many plant species, its functional characteristics in sour jujube have not yet been systematically investigated. Methods: In this study, using Arabidopsis thaliana as a reference, we identified NHX genes in sour jujube through genome-wide analysis and molecular approaches, and systematically analyzed their phylogenetic relationships, chromosomal locations, conserved motifs, gene structures, cis-regulatory elements in promoter regions, and expression patterns under abiotic stresses, particularly salt stress. Results: The results revealed the presence of eight NHX genes distributed across six chromosomes in sour jujube, which were classified into three subfamilies: Vac-class, Endo-class, and PM-class. Members within the same evolutionary clade exhibited high structural conservation in motif composition and gene architecture. Except for the PM-class, all other clades contained amiloride-binding sites (FF(I/L)(Y/F)LFLLPPI). Analysis of cis-regulatory elements indicated that the promoter regions of these genes were enriched with elements related to defense responses, stress adaptation, and phytohormone signaling, further supporting their role in plant environmental adaptation. Additionally, the qRT-PCR analysis showed that most of the ZjNHX genes in both roots and leaves are up-regulated by salt. Notably, ZjNHX1 expression in roots increased approximately 40-fold within 3 h, whereas ZjNHX2 and ZjNHX3 were strongly induced in leaves under prolonged salt exposure. Conclusions: Taken together, this work gives a detailed overview of the ZjNHX genes and their important roles in response to salt stress in sour jujube. Our findings also provide a foundation for further functional characterization of this gene family.