Water stress and sulfur (S) deficiency are two constraints increasingly faced by crops due to climate change and low-input agricultural practices. To investigate their interaction in the grain legume pea (Pisum sativum), sulfate was depleted at the mid-vegetative stage and a moderate 9-d water stress period was imposed during the early reproductive phase. The combination of the stresses impeded reproductive processes in a synergistic manner, reducing seed weight and seed number, and inducing seed abortion, which highlighted the paramount importance of sulfur for maintaining seed yield components under water stress. On the other hand, the moderate water stress mitigated the negative effect of sulfur deficiency on the accumulation of S-rich globulins (11S) in seeds, probably due to a lower seed sink strength for nitrogen, enabling a readjustment of the ratio of S-poor (7S) to 11S globulins. Transcriptome analysis of developing seeds at the end of the combined stress period indicated that similar biological processes were regulated in response to sulfur deficiency and to the combined stress, but that the extent of the transcriptional regulation was greater under sulfur deficiency. Seeds from plants subjected to the combined stresses showed a specific up-regulation of a set of transcription factor and SUMO ligase genes, indicating the establishment of unique regulatory processes when sulfur deficiency is combined with water stress.

Pea is a legume crop producing protein-rich seeds and is increasingly in demand for human consumption and animal feed. The aim of this study was to explore the proteome of developing pea seeds at three key stages covering embryogenesis, the transition to seed-filling, and the beginning of storage-protein synthesis, and to investigate how the proteome was influenced by S deficiency and water stress, applied either separately or combined. Of the 3184 proteins quantified by shotgun proteomics, 2473 accumulated at particular stages, thus providing insights into the proteome dynamics at these stages. Differential analyses in response to the stresses and inference of a protein network using the whole proteomics dataset identified a cluster of antioxidant proteins (including a glutathione S-transferase, a methionine sulfoxide reductase, and a thioredoxin) possibly involved in maintaining redox homeostasis during early seed development and preventing cellular damage under stress conditions. Integration of the proteomics data with previously obtained transcriptomics data at the transition to seed-filling revealed the transcriptional events associated with the accumulation of the stress-regulated antioxidant proteins. This transcriptional defense response involves genes of sulfate homeostasis and assimilation, thus providing candidates for targeted studies aimed at dissecting the signaling cascade linking S metabolism to antioxidant processes in developing seeds.