GhCOMT33D modulates melatonin synthesis, impacting plant response to Cd²⁺ in cotton via ROS

Caffeic acid-3-O-methyltransferase (COMT) serves as the final pivotal enzyme in melatonin biosynthesis and plays a crucial role in governing the synthesis of melatonin in plants. This research used bioinformatics to analyze the phylogenetic relationships, gene structure, and promoter cis-acting elements of the upland cotton COMT gene family members， which it identified as the key gene GhCOMT33D to promote melatonin synthesis and responding to Cd²⁺ stress. After silencing GhCOMT33D through virus-induced gene silencing (VIGS), cotton seedlings showed less resistance to Cd²⁺ stress. Under Cd²⁺ stress, the melatonin content in the silenced plants significantly decreased, while ROS, MDA, and proline accumulated in the plant cells. The stomatal aperture of the leaves was reduced, hindering normal photosynthesis, leading to cotton leaves withering and yellowing, and epidermal cells becoming twisted and deformed, with a large number of gaps appearing. The non-silenced plants had a significantly higher melatonin content and were in better condition, providing important evidence for further research on how plant melatonin enhances the Cd²⁺ resistance of cotton and its regulatory mechanisms.