Functional analysis of OsST41 regulating salt tolerance in rice seedlings

Rice is the most widely cultivated grain crop in China, and its yield is highly susceptible to soil salinization. In our previous study, a candidate gene associated with salt tolerance in rice, <italic>OsST41</italic>, was identified through a genome-wide association study (GWAS). In the present study, we cloned the <italic>OsST41</italic> (<italic>Os07g0598200</italic>) gene. Bioinformatic analysis showed that the gene has a full-length coding sequence of 747 bp and encodes a 248-amino-acid protein containing a conserved F-box domain. The OsST41 protein shares homology with the EDL3 protein in <italic>Arabidopsis thaliana</italic>, which is known to be involved in the regulation of abiotic stress responses. Subcellular localization analysis revealed that OsST41 is localized in the nucleus. qRT-PCR analysis indicated that <italic>OsST41</italic> is expressed at all developmental stages of rice, and its expression in leaves is significantly upregulated under salt stress at the seedling stage. To investigate the function of <italic>OsST41</italic> in salt tolerance, we generated a knockout mutant using CRISPR/Cas9 gene-editing technology. The salt tolerance of the <italic>OsST41</italic> knockout mutant was significantly reduced, and the Na⁺/K⁺ under salt stress was markedly higher than that of the wild type. Physiological measurements showed that, under salt stress, the mutant had significantly higher malondialdehyde (MDA) content and lower activities of superoxide dismutase (SOD), catalase (CAT), and proline (Pro) content compared to the wild type. Furthermore, qRT-PCR analysis revealed that the expression levels of antioxidant-related genes <italic>OsALDH22A1</italic> and <italic>OsGPX5</italic> were significantly downregulated in the mutant. These results suggest that the antioxidant defense system is impaired in the mutant, leading to reduced reactive oxygen species (ROS) scavenging capacity. Collectively, our findings provide preliminary evidence that <italic>OsST41</italic> plays a positive regulatory role in rice salt tolerance, laying a foundation for further exploration of its molecular mechanisms in response to salt stress.