DNA Methylation-Activated LaCOMT1 Expression Promotes Cluster Root Formation of White Lupin via a Mechanism Involving the Melatonin Synthesis.

White lupin exhibits remarkable adaptability to phosphorus (P)-deficient soil through the development of cluster roots (CR), thereby enhancing P use sufficiency. Despite its crucial role, the underlying mechanism governing CR formation remains elusive. Here, we reveal an elevated DNA methylation level through whole-genome bisulfite sequencing in CR in response to P deficiency, particularly in gene and flanking regions, suggesting a responsive epigenetic mechanism. To further investigate the potential involvement of epigenetic remodelling, we treated lupin plants with the DNA methyltransferase (DNMT) inhibitor 5-azacytidine, which led to a disruption of total DNMT activity and impaired CR formation under phosphorus-deficient conditions. Integrated analysis of methylome and RNA-Seq highlights the methylation of CAFFEIC ACID O-METHYLTRANSFERASE 1 (COMT1), a key enzyme in melatonin synthesis, as pivotal for promoting CR formation in white lupin. Functional validation through overexpression or gene silencing of LaCOMT1 in transgenic lupin roots confirms the positive impact of LaCOMT1 on CR formation. Furthermore, melatonin application directly increases CR numbers, indicating the role of methylation-activated LaCOMT1 in promoting CR formation via melatonin synthesis. Those findings provide insights into the epigenomic landscape of white lupin, establishing a direct genetic link between epigenetic mechanisms and P-deficiency-induced CR formation.