Studies On Regulatory Mechanism Of PIF5 In Dark-induced Senescence Of Arabidopsis Seedlings

Senescence is the last stage of plant or organ development. It can bring some disadvantages for grain output and reduce yeild. Phytochrome-interacting factors (PIFs) are transcription factors and play an important role in the process of plant growth and development. They regulate seed germination, hypocotyl elongation, chlorophyll synthesis and so on. But the relationship between PIFs and senescence in plants remains unclear. Cytokinins, as a type of phytohormones, play a critical role in plant growth and development, including regulating leaf senescence. Nitric oxide (NO) is a ubiquitous messenger and effective molecule in plants and other organisms. Evidence suggested that NO participates in a series of key physiological processes, such as leaf and fruit senescence. Because delayed senescence is important in increasing grain output, research on plant senescence has become a demand in national major strategic development. In the present study, using model plant Arabidopsis thaliana as materials, the roles of PIF5 in dark-induced senescence were investigated by physiological and molecular biology methods. At the same time, we investigated if 6-BA or NO can also interact with PIF5 in regulating the dark-induced senescence. The results showed that:(1) In dark-induced senescence, the chlorophyll content in wild type (Col-0) was higher than PIF5OX, but lower than the mutant pif5-2. Col-0 exhibited stronger stress tolerance than PIF5OX, but weaker than pif5-2. For the senescence marker gene-SENESCENCE-ASSOCIATED GENE (SAG 12), its relative expresssion was higher in Col-0 than pif5-2, and in PIF50X the expression was the highest. The two key genes in chlorophyll degradation, STAY GEEEN (SGR) and NON-YELLOW COLORING-1{NYC-1), showed similar characteristics as SAG12 did. These results suggested that PIF5 palys a positive regulatory role in dark-induced senescence.(2) Exogenous 6-BA can prevent the decrease of chlorophyll content in dark-induced senescence in Col-0, PIF5OX, pif5-2 seedlings. In addition, the MDA content, H2O2 content, and Ion leakage were all reduced by 6-BA treatment compared to the control. The expression of SAG12, SGR, and NYC-1 was also decreased compared to the control. PIF5OX had the most obvious decrease, whereas pif5-2 had little changes. The results suggested that the remission of 6-BA in dark-induced senescence is partially dependent on PIF5.(3) Different concentrations of SNP had different effects in dark-induced senescence in Col-0, PIF5OX, pif5-2 seedlings. For example, the chlorophyll content was restored by 50 μM SNP, and the stress resistance was also incresaed. The expression of SAG12, SGR, and NYC-1 was also lower than the control. When 200 μM SNP was added, an opposite result was observed. The results demonstrated that NO has a dose effect. At low concentrations, it can act as a signal molecule and has a positive function in reducing the dark-induced senescence, which is regulated by PIF5. But at high concentrations, it has a negative function. We proposed that excess NO may generate more O2- which is harmful to plants.(4) The expression of PIF5 was examined in control groups and 6-BA-or 50μM SNP-treated groups. Results showed that the PIF5 expression in experimental groups was lower than the control groups. This further explained why 6-BA and SNP can reduce the dark-induced senescence. But the mechanism of how the gene PIF5 get down still needs to be further explored.