The Progress And Regulation Of Leaf Coloration And Senescence In Ginkgo Biloba L.

Ginkgo biloba is one of the world famous relic species,known as a "living fossil".It not only has special landscape ornamental value because of the unique leaf shape and yellow coloration in autumn,but also has pharmaceutical value,edible value and timber value.Leaf senescence is a process in which plants degenerate,decline and eventually die with age at the organ level,mainly characterized by leaf yellowing,declining photo synthetic activity,destruction of cell structure,degradation of macromolecules,transport of nutrients and changes in gene expression level and so on.In this study,using the leaves of G.biloba during autumn coloration as experimental materials,we investigated morphological structure,pigment content and other related physiological indexes to clarify the physiological mechanism of leaf coloration and senescence in G.biloba.Using the RNA-seq of leaves during senescence,we further analyzed the transcriptional regulatory mechanism of leaf coloration and senescence by molecular and bioinformatics method.The main results are as follows.?1?The morphological and SEM observations of leaves during coloration showed that the leaves gradually turned yellow from October to the end of November and completely turned golden,with sunken stomata and deformed epidermal cells.The leaf color parameters L*,a*and b*decreased firstly and then increased,reaching the highest value at the end of November.?2?Physiological determination of pigment contents showed that the contents of chlorophyll and carotenoid increased firstly and then decreased.The chlorophyll content reached the maximum in September and the carotenoid content reached the maximum in early October.From November,the carotenoid content was higher than the total chlorophyll content,and the Car/Chl ratio increased sharply.Among the components of carotenoids,lutein,luteoxanthin and 6-carotene decreased in the late period of senescence.As for the flavonoids,the contents of quercetin,kaempferol and isorhamnetin were increased in the late period of senescence.?3?The soluble sugar content and soluble protein content during leaf senescence showed a trend of 'increasing first and then decreasing',and reached the highest level at the end of October.H2O2 increased continuously from September,while SOD activity began to decrease in September.?4?By adjusting soil acidity and applying mineral elements,we found that slight acidic soil,Fe and Mn treatment can effectively inhibit chlorophyll degradation.The Car/Chl ratio of slight acidic soil,Fe and Mn treatment were significantly lower than the control,indicating a certain delaying effect on leaf color transition and aging of G.biloba.?5?The transcriptome sequencing of leaves during senescence was carried out based on the Illumina HiSeqTM 4000 platform.In chlorophyll biosynthesis pathway,some key genes such as a CRD gene Gb 17158 and a POR gene Gb₂9310,were significantly down-regulated during leaf coloration,while a NYC gene Gb 29139 involved in chlorophyll degradation was significantly up-regulated.In carotenoid biosynthesis pathway,a LCYB gene Gb 24861 and a VDE gene Gb 32110 were significantly down-regulated,while a crtZ gene Gb₄0633 was up-regulated.In flavonoid synthesis pathway,3 FLS genes involved in quercetin and kaempferol synthesis showed up-regulated expression trend.qRT-PCR results further verified the expression trends of three chlorophyll anabolism genes and three carotenoid anabolism genes.?6?Among the major antioxidant enzyme-related genes,2 SOD genes?Gb₁8839,Gb₂8365?and 6 POD genes?Gb₀8270,Gb₁0278,Gb₁6792,Gb₂7511,Gb₃3388,and Gb₃8604?were identified to be down regulated during leaf coloration and senescence.In addition,some enzyme related genes such as CAT and GST were down regulated.Further qRT-PCR results of the expression patterns of 7 antioxidant genes indicate that the ability of antioxidant enzymes to remove reactive oxygen species decreased during leaf senescence.?7?Among hormone synthesis and signaling pathways,two NCED genes synthesized by Xanthoxin in the ABA synthesis pathway,PP2C and ABF genes involved in ABA signal transduction were up-regulated.Genes involved in JA synthesis,AOS,OPR and other genes were mostly up-regulated,and the JAZ genes involved in JA signal transduction were also mostly up regulated.qRT-PCR validation of the expression patterns of ABF gene and JAZ gene indicate that these genes may be involved in response to ABA and JA in the aging process of leaves.?8?A total of 561 differentially expressed transcription factors were identified,among which 14 NAC transcription factor family genes and 16 WRKY transcription factor family genes were up-regulated during leaf senescence.qRT-PCR results validated that 2 WRKY57 genes were significantly up-regulated during leaf senescence.?9?Furthermore,exogenous stress treatments?ABA,MeJA and UV-B?were used to induce premature senescence of G.biloba seedlings,and the result showed that the plant displayed an obvious yellowing and senescence features after treatments.Further qRT-PCR results showed the expressional patterns of 4 pigment anabolism genes,4 antioxidant enzyme related genes,2 hormone signal transduction related genes and 2 transcription factors.The results suggest that they were involved in the process of coloration and senescence of G.biloba leaves.