The Roles Of Polyamine Oxidative Degradation In The Process Of Lettuce Seed Germination

Spermine (Spm), spermidine (Spd) and their diamine obligate precursor putrescine (Put) are micro molecular aliphatic polycations that are ubiquitous in all plant cells. They have been proposed to be closely associated with plant development, morphogenesis and responses to diverse stresses. The recent research indicated that besides being only a means of degrading cellular polyamines, thus contributing to polyamine homeostasis, polyamine degradation participates in important physiological processes through their reaction products. The production of hydrogen peroxide (H₂O₂) deriving from polyamine oxidation has been correlated with cell wall maturation and lignification during development as well as with wound-healing and cell wall reinforcement during pathogen invasion. As a signal molecule, H₂O₂ can mediate programmed cell death, hypersensitive response and defense gene expression. However, whether the polyamine oxidative degradation is directly involved in the seeds germination? What kind of role which H₂O₂ can play in the process of germination? These questions are far from clear. In this study, we choose a normal breed of lettuce (Lactuca sativa L.) seed (Guasihong) as an experimental material to investigate the role of polyamine oxidative degradation in the process of lettuce seed germination. The results are as follows:Treatments with exogenous polyamines at low concentrations promoted early germination of lettuce seed. 0.5 mM Spm was more effective than Put, but less than Spd. The detection of time course of seed germination showed that polyamine soaking could also significantly promote the early germination of lettuce seed, but this effect disappeared after 48 h. Consistent treatment with 1.0 mM Cyclohexylamine (CHA, spermidine synthase inhibitor) had a strong inhibitory effect on the lettuce seed germination. The results from the determination of polyamine effects on lettuce seedling development showed that 0.5 mM Spd could significantly promote root growth as its length increased. Unlike the root development, polyamines inhibited the height of plant. However CHA could promote root development and inhibit plant height. These results suggested that polyamines are involved in the lettuce seed germination and seedling development.The changes of DAO, PAO activities and endogenous H₂O₂ content after treatment with Spd showed that PAO and DAO activities were significantly promoted by Spd, and reached peaks at 24 h and 36 h, respectively. Compared with the control, endogenous H₂O₂ content was also promoted by Spd treatment and had an extremely significant difference after 24 h. Thus, it is proposed that Spd-promoted seed germination may relate to the promotion of polyamine oxidase activity and increase in H₂O₂ content. In order to observe the effect of polyamine degradation on seed germination, aminoguanidine (AG, a specific inhibitor of polyamine oxidase) was employed. The results showed that seed germination was reduced after AG treatment comcomitantly with its inhibitory effect on DAO and PAO activities and resulting in decrease of H₂O₂ content. It was interesting that simultaneous treatment with 0.5mM Spd and 0.1 mM AG could strongly inhibit the Spd-induced increase in DAO and PAO activities and reduce H₂O₂ content, then inhibited the Spd-induced seed germination. In addition, we found that H₂O₂ scavenger (N,N-dimethyl-thiourea: DMTU) treatment strongly inhibited seed germination by reducing H₂O₂ content. 2.0 mM DMTU eliminated the promotive effect of Spd on seed germination and had a similar role with AG.After treatment with Spd and AG, changes in endogenous polyamines in lettuce seeds were determined. These results showed that Put had the highest content, Spd came next and Spm was the lowest. During the seed germination, endogenous Put reduced sharply in the first 24 h, then reduced slowly in 24-60 h, while Spd decreased slowly in the early stage of germination, and then sharply declined after 48 h. However, Spm content was low but slightly increased, and the total polyamine gradually decreased. AG treatment could significantly increased endogenous polyamines, especially Put and Spd. The growth of lettuce seedling roots which were treated by AG and DMTU had significantly changes. Treatment with 1.0 mM AG significantly decreased the length of roots, and inhibited the promotion of 0.5 mM Spd. While 2.0 mM DMTU also inhibited the growth of root, and significantly inhibited the height of lettuce seedlings.In this study, we used exogenous polyamines, polyamine biosynthesis inhibitor, specific inhibitor of polyamine oxidative degradation, and H₂O₂ scavenger to determine the activities of DAO and PAO, H₂O₂ content, endogenous polyamines and seeds germination rate in the germination lettuce seeds. The results obtained here indicated that polyamine oxidative degradation, especially degradation products of polyamine oxidation of H₂O₂, may play an important role in the lettuce seed germination.