Genome-wide identification and expression analysis of proline synthesis and catabolism genes in kiwifruit: exploring the role of AcP5CS1 in salt tolerance

The regulation of proline metabolism is critical for enhancing plant stress tolerance by promoting proline accumulation under abiotic stress conditions. Key enzymes in this pathway include Δ1-pyrroline-5-carboxylate synthase (P5CS), pyrroline-5-carboxylate reductase (P5CR), ornithine δ-aminotransferase (δ-OAT), proline dehydrogenase (PDH), and pyrroline-5-carboxylate dehydrogenase (P5CDH). Despite their importance, comprehensive identification and characterization of these gene families in kiwifruit (<i>Actinidia chinensis</i>) remain unexplored. This study identified two <i>AcP5CSs</i>, one <i>AcP5CR</i>, one <i>AcOAT</i>, three <i>AcPDHs</i>, and one <i>AcP5CDH</i> within the kiwifruit genome. This research comprehensively examined phylogenetic tree, gene structure, motif analysis, cis-regulatory elements and chromosomal distributions analysis, as well as expression profiles under abiotic stresses and hormonal stress. Under salt stress, transcriptional profiling showed marked upregulation of <i>AcP5CS1, AcP5CR</i>, and <i>AcOAT</i>, while <i>AcP5CDH</i> was significantly suppressed, as confirmed by qRT-PCR. Functional analysis demonstrated that <i>AcP5CS1</i> overexpression in Arabidopsis significantly enhanced salt tolerance. The correlation results indicated a strong association between the <i>AcNAC30</i> transcription factors (TFs) and the expression of <i>AcP5CS1</i>. Mechanistic studies using dual-luciferase and electrophoretic mobility shift assays (EMSA) confirmed that <i>AcNAC30</i> directly binds to the <i>AcP5CS1</i> promoter. Therefore, we speculated that <i>AcNAC30</i> likely enhances proline accumulation under salt stress by upregulating the expression of proline metabolic pathway genes. These findings elucidate the genomic architecture of proline metabolic genes in kiwifruit and establish their pivotal role in mediating abiotic stress tolerance.