Genome-Wide Identification of the BhKCS Gene Family in Wax Gourd (Benincasa hispida) and Stress-Responsive Expression of BhKCS8/9/14/15

Abstract

The β-ketoacyl-CoA synthase (KCS) family catalyzes the rate-limiting step in very-long-chain fatty acid (VLCFA) synthesis and plays critical roles in cuticular wax formation and abiotic stress responses. However, the KCS gene family in wax gourd (Benincasa hispida) has remained uncharacterized. In this study, 27 BhKCS genes were identified genome-wide and unevenly distributed across ten chromosomes, with a dense cluster on chromosome 1. Phylogenetic analysis classified the BhKCS proteins into five groups together with Arabidopsis KCSs. Conserved motif and exon-intron structure analyses revealed that most members in Cluster I contain no introns or only one intron, whereas BhKCS24–27 genes possess 14 exons. Promoter cis-element analysis showed that stress-responsive elements were the most abundant, followed by hormone-responsive and light-responsive elements, whereas development-related elements were relatively few. Subcellular localization showed that all four BhKCS fusion proteins localized to the cell membrane and co-localized with the plasma-membrane marker. Heterologous expression in yeast demonstrated that overexpression of BhKCS8 and BhKCS9 maintained yeast growth under osmotic, salt and acid stress conditions, whereas overexpression of BhKCS14 and BhKCS15 suppressed yeast growth under the same conditions, suggesting a functional dichotomy within the family in the yeast system. Scanning electron microscopy revealed that the powder wax gourd epidermis had a more continuous wax coating, whereas the green wax gourd epidermis showed a less continuous coating with larger areas of exposed epidermal surface. Expression profiling under cold, salt and drought stresses showed that several BhKCS genes exhibited variety-specific expression patterns between the green and powder wax gourd types, implying potential differences in their stress responsiveness. This study provides the first systematic characterization of the BhKCS gene family in wax gourd and identifies candidate genes for further functional validation.