| Peach（Prunus persica L.）is a typical respiratory climacteric fruit and is prone to spoilage in room temperature storage.Low temperature can inhibit respiration and other metabolic activities in peach fruit.But peach fruit can easily produce chilling injury symptoms under long-term low-temperature storage,resulting in excessive accumulation of reactive oxygen species（ROS）,leading to browning,lignification,and flavor loss.ROS induces oxidative damage and DNA oxidative damage in cells at high concentrations.N-acetylcysteine（NAC）can inhibit the production of ROS.Nitric oxide（NO）has also been shown to activate the antioxidant system and reduce the production of ROS in peach fruit during storage.Base excision repair（BER）repaired most of the DNA damage produced by ROS.In the BER pathway,replication DNA polymerases such as DNA polymeraseε（Polε）are required to complete the replicative extension of DNA.However,the effect of NO on BER in postharvest fruit cells in the presence of excess or partial removal of ROS and the role of Pp Polεare unknown.In this study,peaches were treated with sodium nitroprusside（SNP）solution,N-acetylcysteine（NAC）solution,a mixture of SNP and NAC,and a mixture of SNP and hydrogen peroxide（H2O2）solution,respectively.The effects of different treatments on the apparent indexes,mitochondrial function,mitochondrial antioxidant system,mitochondrial DNA damage,mitochondrial DNA BER pathway,and Polεin the BER pathway of peach fruit during cold storage at 0°C were studied.The function of peach Polεwas verified by overexpressing peach Polεwith Agrobacterium tumefaciens transient expression system.The conclusions are as follows:（1）SNP+NAC treatment could achieve the best results in reducing oxidative stress and decreasing ROS content in mitochondria of peach fruit by regulating the activities of antioxidant enzymes（SOD）,peroxidase（POD）,and catalase（CAT）.SNP and NAC combination treatment reduced mitochondrial respiratory oxygen consumption,increased mitochondrial respiratory control rate,maintained lower mitochondrial membrane potential,and reduced mitochondrial permeability transition pore（MPTP）.Therefore,with partial removal of reactive oxygen species,NO could better maintain stable mitochondrial function during storage to prolong the storage period of postharvest fruit.（2）SNP+NAC treatment reduced the mitochondrial 8-hydroxyguanosine（8-OHd G）level and decreased the number of apurinic/apyrimidinic sites（AP site）.In addition,SNP and NAC combination treatment up-regulated the activity of enzymes and gene expression levels associated with the BER pathway,which contributed to the ability of mt DNA replication.Therefore,SNP+NAC treatment enhanced the BER pathway,improved the repair efficiency of mt DNA,and helped maintain the integrity of mt DNA.（3）In this study,the Pp Polεwas cloned by RT-PCR,and the sequence of the CDS region was 1590 bp,encoding 529 amino acids,a hydrophilic protein.Secondary structure prediction of protein revealed that theα-helix and irregular helix in the peach fruit Pp Polεare the main structural elements in the overall protein structure.Pp Polεfrom peach fruit was transiently transformed in tomato and peach,and the results showed that overexpression of Pp Polεcould enhance the transcript levels of genes related to the BER pathway,inhibited the reduction of mitochondrial membrane potential and fluidity,and maintained mitochondrial function in peach fruit.Overexpression of Pp Polεcould reduce the MDA content,increase total phenol and total flavonoid content,increase fruit antioxidant capacity through enzymatic antioxidation,maintain fruit quality and reduce oxidative damage in peach fruit.Conclusion:The combination of SNP and NAC treatment significantly improved the scavenging ability of reactive oxygen species in mitochondria of frozen peach fruit,reduced oxidative damage in peach fruit during low temperature storage.SNP and NAC combination treatment regulated the activity of key enzymes and gene transcription levels in the BER pathway and reduced mt DNA damage.Overexpression of Pp Polεup-regulated the expression of BER-related genes in tomato and peach fruit and improved storage quality of peach fruit. |
