Cotton 4-coumarate-CoA ligase 3 enhanced plant resistance to Verticillium dahliae by promoting JA signaling mediated vascular lignification and metabolic flux.

Lignins and their antimicrobial related polymers cooperatively enhance plant resistance to pathogens. Several isoforms of 4-coumarate: Coenzyme A ligases (4CLs) have been identified as indispensable enzymes involved in lignin and flavonoids biosynthetic pathways. However, their roles in plant-pathogen interaction are still poorly understood. This study uncovers the role of Gh4CL3 in cotton resistance to the vascular pathogen Verticillium dahliae. Cotton 4CL3-CRISPR/Cas9 mutant (CR4cl) exhibited high susceptibility to V. dahliae. This susceptibility was most probably due to the reduction in total lignin content and the biosynthesis of several phenolic metabolites, e.g. rutin, catechin, scopoletin glucoside and chlorogenic acid, along with Jasmonic acid (JA) attenuation. These changes were coupled with a significant reduction in 4CL activity toward p-coumaric acid substrate. Likely, recombinant Gh4CL3 could specifically catalyze p-coumaric acid to form p-coumaroyl-CoA. Thus, overexpression of Gh4CL3 (OE4CL) showed increasing 4CL activity that augmented phenolic precursors, cinnamic, p-coumaric and sinapic acids, channeling into lignin and flavonoids biosyntheses and enhanced resistance to V. dahliae. Besides, Gh4CL3 overexpression activated JA signaling that instantly stimulated lignin deposition and metabolic flux in response to pathogen which all established efficient plant defense response system, and inhibited V. dahliae mycelium growth. Our results propose that Gh4CL3 acts as a positive regulator for cotton resistance against V. dahliae by promoting JA signaling mediated enhanced-cell wall rigidity and metabolic flux.