Whole genome identification and analysis of cotton(Gossypium hirsutum L.) pyruvate kinase gene family and functional analysis of GhPK5 gene

Background Pyruvate kinase (PK) is a key rate-limiting enzyme in the glycolytic pathway, playing a central role in plant energy metabolism, growth, development, and stress responses. Although the PK gene family has been characterized in several model plants, systematic identification and functional analysis of PK genes in cotton ( Gossypium hirsutum L.) remain limited. Results In this study, we identified 41 PK genes in the upland cotton genome and classified them into two subfamilies (PKc and PKp) based on phylogenetic analysis. Chromosomal distribution analysis revealed that these genes are unevenly distributed across 19 chromosomes, with segmental duplication serving as the predominant expansion mechanism. Gene structure and conserved motif analyses demonstrated that members within the same subfamily share similar exon-intron organizations and motif compositions. Cis-acting element analysis indicated that GhPK promoters are enriched with stress-responsive and hormone-responsive elements. Transcriptome profiling showed diverse expression patterns of GhPK genes across different tissues, with GhPK5 exhibiting high and broad expression. Under salt, drought, heat, and cold stresses, GhPK5 displayed a strong and sustained induction, particularly under salt and combined stress conditions. Virus-induced gene silencing (VIGS) of GhPK5 resulted in increased SOD, POD, CAT activities and MDA content under salt stress, along with impaired pyruvate accumulation, suggesting that GhPK5 plays a positive role in cotton salt tolerance. Conclusions Our genome-wide characterization of the cotton PK gene family provides a foundational understanding of their evolutionary and functional diversity. GhPK5 was identified as a candidate gene positively regulating salt stress response, offering a potential target for molecular breeding aimed at improving stress tolerance in cotton.