Résumé
In 1962, a landmark study by Murashige & Skoog (MS) revealed the dose-dependent effect of nitrogen (N) on tobacco calli biomass was enhanced by increasing the doses of phosphorus (P) and potassium (K). Their study established MS media as a gold standard for plant growth in tissue culture which has been used in plant science research for decades. Now in the genomic era, we seek to understand the molecular basis for this genetic integration of N, P, and K (NPK) signals and the phenotypic outcomes they affect. As overuse of N fertilizer is costly – both monetarily and environmentally - understanding how P and K can enhance N-dose dependent growth responses could have important agricultural implications. To uncover the molecular basis of NPK signaling interactions, we repeated the Murashige and Skoog experiments in Arabidopsis thaliana. Indeed, we found that increasing P and K doses nearly doubles shoot biomass, specifically under intermediate N-dose conditions (5-20mM N). We then collected transcriptome and phenotype trait data from Col-0 plants grown under an NPK dose matrix; a continuous N-dose gradient (6 doses) at 4 PK conditions (Low and High, P and K doses: PK, pK, Pk, pk). A DESeq2-based linear model simplification analysis of the RNA-seq data enabled us to characterize significant gene expression changes in response to distinct combinations of NPK doses. We found that different nutrient-interaction-responsive gene lists (N:P, N:K, or N:P:K) are enriched for distinct biological processes related to plant growth. For example the N:K-responsive gene list is enriched for photosynthesis related genes, while the N:P-responsive gene list is enriched for drought response genes. N:P:K-responsive genes are enriched for rRNA processing, transcriptional regulation, and circadian rhythm, revealing how the integration of NPK nutrients influences transcription and metabolism. To validate regulators of these NPK signaling interactions, we conducted mutant analysis of three nutrient interaction-responsive genes and found significant changes in Arabidopsis root and/or shoot growth under specific NPK combinations. To uncover the underlying gene regulatory networks, we conducted RNA-seq analysis of one root N:P:K-responsive transcription factor (TF) mutant, compared to wild-type Col-0 plants. DESeq2 differential expression analysis revealed that thousands of root nutrient-responsive genes are dysregulated in specific NPK conditions when the TF is mutated. Lastly, using weighted gene correlation network analysis (WGCNA), we showed that NPK interaction-responsive genes are correlated to biomass and enriched for growth related processes. For example, N:K-responsive photosynthesis genes are positively correlated to biomass, while N:K-responsive stress signaling genes are negatively correlated to biomass. Co-expressed N:P:K responsive genes are correlated to lateral root branching density and are enriched for DNA replication and gene expression regulation. Our study opens a window into the molecular mechanisms by which Arabidopsis can sense and respond to distinct combinations of NPK doses, which in turn lead to dramatic changes in plant growth and development.