Abstract
BACKGROUND: Breeding for polyploidy in Brassica campestris L. has been employed to enhance agronomic traits. Polyploid plants exhibit improved productivity due to the 'gigas' effect driven by chromosome doubling. In our previous study, autotetraploids were induced in Pakchoi (Brassica campestris L. ssp. chinensis) via colchicine, resulting in two distinct autotetraploid genotypes: gigantic genotype 'P4x-3' with high-yield potential and dwarf genotype 'P4x-7' with low-yield traits in comparison to their diploid progenitor, 'P2x'. However, the molecular mechanisms underlying this phenotypic divergence between the two Pakchoi autotetraploids and their diploid progenitor remain poorly understood. RESULTS: Significant phenotypic differences were observed between the two autotetraploid genotypes and their diploid progenitor. The per plant fresh weight of 'P4x-3' was significantly higher than that of 'P2x' due to its higher leaf area, leaf width, leaf thickness, petiole width, and petiole thickness. 'P4x-7' showed a significantly lower per plant fresh weight than 'P2x' due to its lower plant height, number of leaves, leaf length, and petiole length. The most differentially expressed genes (DEGs) were identified in 'P4x-7 vs. P2x' with 2337 upregulated and 2067 downregulated DEGs. Gene Ontology enrichment analysis showed that DEGs in all three comparisons ('P4x-3 vs. P2x', 'P4x-7 vs. P2x', and 'P4x-3 vs. P4x-7') were primarily enriched in 'response to stimuli' and 'primary metabolic processes'. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis identified three significantly enriched pathways: 'plant hormone signal transduction', 'MAPK signaling pathway - plant', and 'plant - pathogen interaction'. Therefore, we hypothesized that crosstalk between auxin and ethylene signaling regulates plant growth, directly influencing phenotypic traits and the formation of the gigantic 'P4x-3' and dwarf 'P4x-7' genotypes. DEGs, including those from the AUX/IAA, SAUR, ETS/ERS, and CaM/CML families, were identified as critical regulators of growth and stimulus responses in Pakchoi autotetraploids and potentially caused morphological variation. Additionally, 638, 655, and 704 differentially accumulated metabolites were identified in 'P4x-3 vs. P2x', 'P4x-7 vs. P2x', and 'P4x-3 vs. P4x-7', respectively. Enrichment analysis of indole-type metabolites among plant hormones revealed that 'tryptophan metabolism' was associated with the most enriched metabolites. L-tryptophan abundance followed the order of 'P4x-3' > 'P2x' > 'P4x-7'. This finding was consistent with the endogenous indole-3-acetic acid (IAA) content. The endogenous ethylene content exhibited an opposite trend to that of IAA. CONCLUSIONS: Chromosome doubling modulates phenotypic variation in Pakchoi by integrating gene expression across signaling and regulatory pathways, with a focus on those governing plant growth and stimulus responses. Auxin and ethylene signaling transduction and metabolism may act as key determinants of morphological divergence in autotetraploid Pakchoi. This study provides novel insights into the mechanisms underlying phenotypic variation in autotetraploid, with implications for optimizing polyploid breeding strategies.