Comparative RNA sequencing analyses on two soybean genotypes with contrasting seed size across seed set, growth, and early maturation stages identified key genes, modules, and regulatory networks controlling seed development.

In plant cells, mitochondria are ideally positioned to sense and balance changes in energy metabolism in response to changing environmental conditions. Retrograde signalling from mitochondria to the nucleus is crucial for adjusting the required transcriptional responses. We show that ANAC017, the master regulator of mitochondrial stress, directly recruits a signalling cascade involving the plant hormones ethylene and auxin as well as the MAP KINASE KINASE (MKK) 9 - MAP KINASE (MPK) 3/6 pathway in Arabidopsis thaliana. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) and overexpression demonstrated that ANAC017 directly regulates several genes of the ethylene and auxin pathways, including MKK9, 1-AMINO-CYCLOPROPANE-1-CARBOXYLATE SYNTHASE 2 and YUCCA 5, in addition to genes encoding transcription factors regulating plant growth and stress responses such as BASIC REGION/LEUCINE ZIPPER MOTIF (bZIP) 60, bZIP53, ANAC081/ATAF2 and RADICAL-INDUCED CELL DEATH1. A time-resolved RNA-seq experiment established that ethylene signalling precedes the stimulation of auxin signalling in the mitochondrial stress response, with a large part of the transcriptional regulation dependent on ETHYLENE INSENSITIVE 3. These results were confirmed by mutant analyses. Our findings identify the molecular components controlled by ANAC017, which integrates the primary stress responses to mitochondrial dysfunction with whole plant growth via the activation of regulatory and partly antagonistic feedback loops.

Flooding causes severe crop losses in many parts of the world. Genetic variation in flooding tolerance exists in many species; however, there are few examples for the identification of tolerance genes and their underlying function. We conducted a genome-wide association study (GWAS) in 387 Arabidopsis (Arabidopsis thaliana) accessions. Plants were subjected to prolonged submergence followed by desubmergence, and seven traits (score, water content, Fv/Fm, and concentrations of nitrate, chlorophyll, protein, and starch) were quantified to characterize their acclimation responses. These traits showed substantial variation across the range of accessions. A total of 35 highly significant single nucleotide polymorphisms (SNPs) were identified across the 20 GWA data sets, pointing to 22 candidate genes, with functions in TCA cycle, DNA modification, and cell division. Detailed functional characterization of one candidate gene, ACONITASE3 (ACO3), was performed. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) showed that a SNP in the ACO3 promoter co-located with the binding site of the master regulator of retrograde signaling ANAC017, while subcellular localisation of a ACO3-YFP fusion protein confirmed a mitochondrial localization during submergence. Analysis of mutant and overexpression lines determined changes in trait parameters that correlated with altered submergence tolerance and were consistent with the GWAS results. Subsequent RNA-seq experiments suggested that impairing ACO3 function increases the sensitivity to submergence by altering ethylene signalling, whereas ACO3 overexpression leads to tolerance by metabolic priming. These results indicate that ACO3 impacts submergence tolerance through integration of carbon and nitrogen metabolism via the mitochondrial TCA cycle and impacts stress signaling during acclimation to stress.