Functional characterization of Gossypium arboreum 3-ketoacyl-CoA synthase-4 in epicuticular wax biosynthesis confers drought and whitefly-mediated viral tolerance in cotton.

Abstract

Overexpression of the Gossypium arboreum 3-ketoacyl-CoA synthase-4 (GaKCS4) gene, a key enzyme involved in very-long-chain fatty acid (VLCFA) elongation, was evaluated for its role in epicuticular wax biosynthesis and stress tolerance in T0 transgenic Gossypium hirsutum plants. A transformation efficiency of 3.8% was achieved through Agrobacterium-mediated genetic transformation. Transgenic plants exhibited significant upregulation of GaKCS4, accompanied by increased expression of the wax biosynthesis-related genes CER3 and KCS6, enhanced epicuticular wax accumulation, and enrichment of VLCFA-derived wax constituents. Increased wax deposition was associated with improved drought tolerance, as evidenced by higher relative leaf water content, reduced transpiration, and sustained photosynthetic activity. Under free-choice bioassays, transgenic plants showed significantly reduced whitefly colonization, lower Cotton Leaf Curl Virus (CLCuV) β-satellite accumulation, and milder disease symptoms compared with non-transgenic controls. Strong negative correlations were observed between wax load and both whitefly population (r = -0.90) and disease incidence (r = -0.98), indicating a close association between enhanced wax deposition and reduced biotic stress. Furthermore, transgenic plants exhibited an approximately 11-fold increase in trichome density (285 vs. 26 trichomes cm-2), which may have contributed to reduced insect preference. Collectively, these findings demonstrate that GaKCS4-mediated activation of the wax biosynthetic pathway enhances both drought tolerance and resistance to whitefly-mediated viral infection, highlighting its potential for developing climate-resilient cotton cultivars.