Multi-Omics Analysis of Reproductive Phenotypes and Seed Nutritional Remodeling in Soybean GmPEAMT1/GmPLMT Double Knockouts

The soybean GmPEAMT1/GmPLMT double-knockout mutant (Dko) was constructed to investigate how the disruption of phospholipid-related genes affects reproductive traits, seed nutrient composition, and associated molecular responses under experimental conditions. Dko plants showed coordinated changes in photosynthetic parameters and reproductive structures: the number of pollen grains and ovules was reduced, whereas pollen and ovule size-related traits were increased, while mature seed number and 100-seed weight were maintained. Developing Dko seeds showed higher protein content and increased levels of 12 out of 18 measured amino acids, particularly alanine and cysteine. Transcriptomic and qRT-PCR analyses further indicated coordinated changes in amino acid, pantothenate, coenzyme A, glycerolipid, and glycerophospholipid-related pathways. Lipidomics provided complementary evidence of changes in phospholipid-associated and other lipid molecular features, whereas total seed lipid content showed no significant difference from the control. Together, these results suggest that GmPEAMT1/GmPLMT disruption was associated with variations in reproductive traits at the developmental stages examined and seed nutrient allocation, with lipid-feature changes representing one component of a broader multi-omic response. The proposed lipid and amino acid metabolic remodeling in GmPEAMT1/GmPLMT double-knockout soybean seeds should be regarded as a working hypothesis because direct metabolic flux, protein abundance, and enzyme activity are not measured.