Mitochondria are the source of reactive oxygen species (ROS) in plant cells and play a central role in the mitochondrial electron transport chain (ETC) and tricarboxylic acid cycle (TCA) cycles; however, ROS production and regulation for seed germination, seedling growth, as well as mitochondrial responses to abiotic stress, are not clear. This study was conducted to obtain basic information on seed germination, embryo mitochondrial antioxidant responses, and protein profile changes in artificial aging in oat seeds (Avena sativa L.) exposed to exogenous nitric oxide (NO) treatment. The results showed that the accumulation of H₂O₂ in mitochondria increased significantly in aged seeds. Artificial aging can lead to a loss of seed vigor, which was shown by a decline in seed germination and the extension of mean germination time (MGT). Seedling growth was also inhibited. Some enzymes, including catalase (CAT), glutathione reductase (GR), dehydroascorbate reductase (DHAR), and monodehydroascorbate reductase (MDHAR), maintained a lower level in the ascorbate-glutathione (AsA-GSH) scavenging system. Proteomic analysis revealed that the expression of some proteins related to the TCA cycle were down-regulated and several enzymes related to mitochondrial ETC were up-regulated. With the application of 0.05 mM NO in aged oat seeds, a protective effect was observed, demonstrated by an improvement in seed vigor and increased H₂O₂ scavenging ability in mitochondria. There were also higher activities of CAT, GR, MDHAR, and DHAR in the AsA-GSH scavenging system, enhanced TCA cycle-related enzymes (malate dehydrogenase, succinate-CoA ligase, fumarate hydratase), and activated alternative pathways, as the cytochrome pathway was inhibited. Therefore, our results indicated that seedling growth and seed germinability could retain a certain level in aged oat seeds, predominantly depending on the lower NO regulation of the TCA cycle and AsA-GSH. Thus, it could be concluded that the application of 0.05 mM NO in aged oat seeds improved seed vigor by enhancing the mitochondrial TCA cycle and activating alternative pathways for improvement.

Seeds lose their viability when they are exposed to high temperature and moisture content (MC) during storage. The expression and metabolism of proteins plays a critical role in seed resistance to heat stress. However, the proteome response to heat stress in oat (Avena sativa) seeds during storage has not been revealed. To understand mechanisms of heat stress acclimation and tolerance in oat seeds, an integrated physiological and comparative proteomic analysis was performed on oat seeds with different MC during heat stress. Oat seeds with 10% and 16% MC were subjected to high temperatures (35, 45 and 50°C) for 24 days and 2 days, respectively, and changes in physiological and biochemical characteristics were analyzed. The results showed that seed vigor decreased significantly with temperature increase from 35 to 50°C. Also, the proline content in 10% MC seeds decreased significantly (p<0.05) whereas that in 16% MC seeds increased significantly (p<0.05) during heat treatment from 35 to 50 °C. There were no significant differences in malondialdehyde content in 10% MC seeds with temperature from 35 to 50°C, but a significant (p<0.05) decline occurred in 16% MC seeds at 45°C. Proteome analysis revealed 21 significantly different proteins, including 19 down-regulated and two up-regulated proteins. The down-regulated proteins, notably six heat shock proteins and two ATP synthases, have important roles in the mobilization of carbohydrates and energy, and in the balance between synthesis and degradation of other proteins during seed deterioration. The up-regulation of argininosuccinate synthase participated in proline biosynthesis at 16% MC, which is important for maintaining reactive oxygen species homeostasis for the resistance of heat stress. In summary, heat-responsive protein species and mitochondrial respiratory metabolism were sensitive to high temperature and MC treatment. These studies provide a new insight into acclimation and tolerance to heat stress in oat seeds.

Flower development, pollination, and fertilization are important stages in the sexual reproduction process of plants; they are also critical steps in the control of seed formation and development. During alfalfa (Medicago sativa L.) seed production, some distinct phenomena such as a low seed setting ratio, serious flower falling, and seed abortion commonly occur. However, the causes of these phenomena are complicated and largely unknown. An understanding of the mechanisms that regulate alfalfa flowering is important in order to increase seed yield. Hence, proteomic technology was used to analyze changes in protein expression during the stages of alfalfa flower development. Flower samples were collected at pre-pollination (S1), pollination (S2), and the post-pollination senescence period (S3). Twenty-four differentially expressed proteins were successfully identified, including 17 down-regulated in pollinated flowers, one up-regulated in pollinated and senesced flowers, and six up-regulated in senesced flowers. The largest proportions of the identified proteins were involved in metabolism, signal transduction, defense response, oxidation reduction, cell death, and programmed cell death (PCD). Their expression profiles demonstrated that energy metabolism, carbohydrate metabolism, and amino acid metabolism provided the nutrient foundation for pollination in alfalfa. Furthermore, there were three proteins involved in multiple metabolic pathways: dual specificity kinase splA-like protein (kinase splALs), carbonic anhydrase (CA), and NADPH: quinone oxidoreductase-like protein (NQOLs). Expression patterns of these proteins indicated that MAPK cascades regulated multiple processes, such as signal transduction, stress response, and cell death. PCD also played an important role in the alfalfa flower developmental process, and regulated both pollination and flower senescence. The current study sheds some light on protein expression profiles during alfalfa flower development and contributes to the understanding of the basic molecular mechanisms during the alfalfa flowering process. These results may offer insight into potential strategies for improving seed yield, quality, and stress tolerance in alfalfa.