Rice melatonin deficiency causes premature leaf senescence via DNA methylation regulation

In a study of DNA methylation changes in melatonin-deficient rice mutants,mutant plants showed premature leaf senescence during grain-filling and reduced grain yield.Melatonin deficiency led to transcriptional reprogramming,especially of genes involved in chlorophyll and carbon metabolism,redox regulation,and transcriptional regulation,during dark-induced leaf senescence.Hypomethylation of mCG and mCHG in the melatonin-deficient rice mutants was associated with the expression change of both protein-coding genes and transposable element-related genes.Changes in gene expression and DNA methylation in the melatonin-deficient mutants were compensated by exogenous application of melatonin.A decreased S-adenosyl-L-methionine level may have contributed to the DNA methylation variations in rice mutants of melatonin deficiency under dark conditions.

Synergistic effects of planting density and nitrogen fertilization on chlorophyll degradation and leaf senescence after silking in maize

Regulating planting density and nitrogen(N)fertilization could delay chlorophyll(Chl)degradation and leaf senescence in maize cultivars.This study measured changes in ear leaf green area(GLA_(ear)),Chl content,the activities of Chl a-degrading enzymes after silking,and the post-silking dry matter accumulation and grain yield under multiple planting densities and N fertilization rates.The dynamic change of GLA_(ear)after silking fitted to the logistic model,and the GLA_(ear) duration and the GLAearat 42 d after silking were affected mainly by the duration of the initial senescence period(T_(1))which was a key factor of the leaf senescence.The average chlorophyllase(CLH)activity was 8.3 times higher than pheophytinase activity and contributed most to the Chl content,indicating that CLH is a key enzyme for degrading Chl a in maize.Increasing density increased the CLH activity and decreased the Chl content,T1,GLAear,and GLA_(ear) duration.Under high density,appropriate N application reduced CLH activity,increased Chl content,prolonged T1,alleviated high-density-induced leaf senescence,and increased post-silking dry matter accumulation and grain yield.

Rice ONAC016 promotes leaf senescence through abscisic acid signaling pathway involving OsNAP

Senescence-induced NAC(senNAC)TFs play a crucial role in senescence during the final stage of leaf development.In this study,we identified a rice senNAC,ONAC016,which functions as a positive regulator of leaf senescence.The expression of ONAC016 increased rapidly in rice leaves during the progression of dark-induced and natural senescence.The onac016-1 knockout mutant showed a delayed leaf yellowing phenotype,whereas the overexpression of ONAC016 accelerated leaf senescence.Notably,ONAC016 expression was upregulated by abscisic acid(ABA),and thus detached leaves of the onac016-1 mutant remained green much longer under ABA treatment.Quantitative RT-PCR analysis showed that ONAC016 upregulates the genes associated with chlorophyll degradation,senescence,and ABA signaling.Yeast one-hybrid and dual-luciferase assays revealed that ONAC016 binds directly to the promoter regions of OsNAP,a key gene involved in chlorophyll degradation and ABA-induced senescence.Taken together,these results suggest that ONAC016 plays an important role in promoting leaf senescence through the ABA signaling pathway involving OsNAP.

Slight shading after anthesis increases photosynthetic productivity and grain yield of winter wheat (Triticum aestivum L.) due to the delaying of leaf senescence

The solar radiation intensity and duration are continuously decreasing in the major wheat planting area of China. As a con- sequence, leaf senescence, photosynthesis, grain filling and thus wheat yield shall be affected by light deficiency. Therefore, two winter wheat （Triticum aestivum L.） cultivars, Tainong 18 （a large-spike cultivar） and Ji＇nan 17 （a multiple-spike cultivar）, were subjected to shading during anthesis and maturity under field condition in 2010-2011 and 2011-2012. Under the slight shading treatment （$1,88% of full sunshine）, leaf senescence was delayed, net photosynthesis rate （Po） and canopy apparent photosynthesis rate （CAP） were improved, and thus thousand-kernel weight （TKW） and grain yield were higher as compared with the control. However, mid and severe shading （S2 andS3, 67 and 35% of full sunshine, respectively） led to negative effects on these traits substantially. Moreover, superoxide dismutase （SOD）, peroxidase （POD） and cat- alase （CAT） activities in flag leaf were significantly greater under slight shading than those in other treatments, while the malondialdehyde （MDA） content was less than that under other treatments. In addition, the multiple-spike cultivar is more tolerant to shading than large-spike cultivar. In conclusion, slight shading after anthesis delayed leaf senescence, enhanced photosynthesis and grain filling, and thus resulted in higher grain yield.

Effects of Aerated Irrigation on Leaf Senescence at Late Growth Stage and Grain Yield of Rice

With the japonica inbred cultivar Xiushui 09, indica hybrid combinations Guodao 6 and Liangyoupeijiu as materials, field experiments were conducted in 2007 and 2008 to study the effects of aerated irrigation on leaf senescence at late growth stage and grain yield of rice. The dissolved oxygen concentration of aerated water evidently increased and decreased at a slow rate. The soil oxidation-reduction potential under aerated irrigation treatment was significantly higher than that of the CK, contributing to significant increases in effective panicles, seed setting rate and grain yield. In addition, the aerated irrigation improved root function, increased superoxide dismutase activity and decreased malondialdehyde content in flag leaves at post-flowering, which delayed leaf senescence process, prolonged leaf functional activity and led to enhanced grain filling.

Exogenous abscisic acid coordinating leaf senescence and transport of assimilates into wheat grains under drought stress by regulating hormones homeostasis

Drought at the grain filling stage of wheat will cause premature leaf senescence, thus leading to considerable loss of wheat yield. Therefore, this paper aims to establish a cultivation technology for strong drought resistance, delayed senescence, and yield improvement based on the analysis of hormones homeostasis obtained by applying chemical control substances. Experiments were conducted with two genotypes of wheat. Four water irrigation treatments were applied to impose the water deficit, including well-watered control treatment(WW), mild water deficit(MiWD), moderate water deficit(MoWD), and severe water deficit(SWD). Exogenous abscisic acid(ABA) was sprayed on the plants at the anthesis stage of the wheat. As a result, exogenous ABA reduced initial senescence rate(r0), total duration of chlorophyll(Chltotal), rapid senescence phase(Chlloss), and the accumulated temperature at an inflection point(M) but improved the persistence phase(Chlper) of flag leaves under all of the four treatments. However, exogenous ABA produced inconsistent effects on photoassimilate relocation and grain weight under different treatments. It produced positive regulatory effects on grain weight under WW, MiWD, and MoWD treatments. On the one hand, spraying ABA during the persistence phase of flag leaves reduced the ratios of zeatin to gibberellin(Z/GA3), spermine to spermidine(Spm/Spd), and salicylic acid to ABA(SA/ABA),which prolonged active photosynthesis by stimulating high level of proline(Pro) and increased the activities of antioxidant enzymes, such as superoxide dismutase(SOD), peroxidase(POD), catalase(CAT), and ascorbate peroxidase(APX). Therefore, drought tolerance was enhanced, and more photosynthetic assimilates were accumulated. On the other hand, the rapid senescence phase and the transport rate of assimilates into grains were accelerated, resulting in higher grain weight, yield, and water use efficiency(WUE). However, under SWD treatment, exogenous ABA improved the ratio of SA/ABA, leading to low Pro content and low antioxidant enzyme activity of flag leaves in the rapid loss phase. Meanwhile,drought resistance declined and the transport duration of assimilates into grains was shortened, thus making photosynthetic assimilates redundant. Therefore, exogenous ABA can lead to the reduction in grain weight, yield, and WUE of wheat under SWD treatment.

Mapping a leaf senescence gene els1 by BSR-Seq in common wheat

Leaf senescence is normally the last stage of plant development. Early senescence of functional leaves significantly reduces the photosynthetic time and efficiency, seriously affecting grain yield and quality in wheat. Discovering genes responsible for early leaf senescence(els) are necessary for developing novel germplasms and cultivars with delayed leaf-senescence through molecular manipulation and marker assisted selection. In this study, we identified an early leaf senescence line M114 in a derivative of a wheat breeding population. Genetic analysis indicated that early leaf senescence in M114 is controlled by a single recessive gene, provisionally designated els1. By applying bulked segregant analysis and RNA-Seq(BSR-Seq), seven polymorphic markers linked to els1 were developed and the gene was located on chromosome arm 2 BS in a 1.5 c M genetic interval between markers WGGB303 and WGGB305. A co-segregating marker, WGGB302, provide a starting point for fine mapping and map-based cloning of els1.

A rice XANTHINE DEHYDROGENASE gene regulates leaf senescence and response to abiotic stresses

Xanthine dehydrogenase, a member of the molybdenum enzyme family, participates in purine metabolism and catalyzes the generation of ureides from xanthine and hypoxanthine. However, the mechanisms by which xanthine dehydrogenase affects rice growth and development are poorly understood. In the present study, we identified a mutant with early leaf senescence and reduced tillering that we named early senescence and less-tillering 1(esl1). Map-based cloning revealed that ESL1 encodes a xanthine dehydrogenase, and it was expressed in all tissues. Chlorophyll content was reduced and chloroplast maldevelopment was severe in the esl1 mutant. Mutation of ESL1 led to decreases in allantoin, allantoate, and ABA contents. Further analysis revealed that the accumulation of reactive oxygen species in esl1 resulted in decreased photosynthesis and impaired chloroplast development, along with increased sensitivity to abscisic acid and abiotic stresses. Ttranscriptome analysis showed that the ESL1 mutation altered the expression of genes involved in the photosynthesis process and reactive oxygen species metabolism.Our results suggest that ESL1 is involved in purine metabolism and the induction of leaf senescence.These findings reveal novel molecular mechanisms of ESL1 gene-mediated plant growth and leaf senescence.

Yield, Leaf Senescence, and Cry1Ac Expression in Response to Removal of Early Fruiting Branches in Transgenic Bt Cotton

Two-year field experiments were conducted at Linqing, Yellow River valley of China, to study the plant response to the removal of early fruiting branches in transgenic Bt （Bacillus thuringiensis） cotton （Gossypium hirsutum L.） from 2003 to 2004. Plants were undamaged and treated by removing two basal fruiting branches （FB） at squaring to form the control and the removal treatment, respectively. The plant height, leaf area （LA）, dry weight of fruiting forms （DWFF）, the number of fruiting nodes （NFN）, photosynthetic （Pn） rate, and levels of leaf chlorophyll （Chl）, N, P, K, and Cry lAc protein in main- stem leaves were measured at a 10- or 20-d interval after FB removal, and the sink/source ratio as indicated by NFN/LA and DWFF/LA was determined. FB removal significantly increased the plant height, LA, and plant biomass in both years. Lint yields were increased 7.5 and 5.2% by removal compared with their controls in 2003 and 2004, respectively. Significant increases in boll size （5.7 and 5.1%） were also observed in removal than in control for both years. Either NFN/LA or DWFF/LA was significantly reduced by removal before 40 d after removal; however, both NFN/LA and DWFF/LA were significantly enhanced by FB removal at 80 d after removal compared to the untreated control. There was no significant difference in fiber quality in the first two harvests between removal and control, but fiber strength and micronarie in the third harvest were significantly improved by FB removal. In terms of leaf Chl, Pn rate, levels of total N, P, and K in late season, leaf senescence was considerably delayed by FB removal. Levels of CrylAc protein in the fully expanded young leaves were considerably higher in FB-excised plants than in control, indicating FB removal enhanced CrylAc expression. It is suggested that the yield and quality improvement with FB removal may be attributed to the increased NFN/LA or DWFF/LA in late season and delayed leaf senescence, respectively. FB removal can be a potential practice incorporated into the intensive cultivation system for enhancing transgenic Bt cotton production.

The Changes of Photosynthetic Properties and Cell Microstructure in Peanut Leaves during Leaf Senescence

The changes of photosynthetic properties and cell microstructure in peanut leaves during leaf senescence were studied with two high-yielding peanut cultivars (cv. Luhuall and Fu8707). The main results showed that during the whole process of leaf growth and senescence, changes in the photosynthesis rate (Pn) and contents of chlorophyll in leaves, could be described with a parabolic function, y = A + Bx + Cx2 (where y refers to the values of the above parameters and x to the days after leaf unfolding). During peanut leaf senescence , the shape of chloroplast changed gradually from long ellipses to circles.The starch globule in chloroplast altered gradually from more and larger sizes to fewer and smaller, but the oil globule from fewer and smaller to more and larger. The grana lamellae varied progressively: from thinness and length to thickness and shortness; from ranking along the long axle direction of chloroplast to disorderly arrangment and finally blurring. At last, the membrane envelope of chloroplast broke, so the inclusion seeped out to the cell and the chloroplast broke up.

AtWRKY75 positively regulates age-triggered leaf senescence through gibberellin pathway

WRKY transcription factors play essential roles during leaf senescence.However,the mechanisms by which they regulate this process remains largely unknown.Here,we identified the transcription factor WRKY75 as a positive regulator during leaf senescence.Mutations of WRKY75 caused a delay in agetriggered leaf senescence,whereas overexpression of WRKY75 markedly accelerated this process.Expression of senescence-associated genes(SAGs)was suppressed in WRKY75 mutants but increased in WRKY75-overexpressing plants.Further analysis demonstrated that WRKY75 directly associates with the promoters of SAG12 and SAG29,to activate their expression.Conversely,GAI and RGL1,two DELLA proteins,can suppress the WRKY75-mediated activation,thereby attenuating SAG expression during leaf senescence.Genetic analyses showed that GAI gain-of-function or RGL1 overexpression can partially rescue the accelerated senescence phenotype caused by WRKY75 overexpression.Furthermore,WRKY75 can positively regulate WRKY45 expression during leaf senescence.Our data thus imply that WRKY75 may positively modulate age-triggered leaf senescence through the gibberellin-mediated signaling pathway。

Effects of 4PU-30 on leaf senescence and degration of protein and nucleic acid in rice

4PU—30[N—phenyl—’N—(2—chloro—4—pyridyl) urea] is a new type of plant growth regulator with cytokinin properties. It has been confirmed to delay rice leaf senescence effectively. In order to elucidate the physiological role of 4PU—30 in delaying senescence, the changes of protein, nucleic acid contents, and the related activities of degradative enzymes were studied. Shanyou 63, an indica hybrid rice was used for this experiment. In the in vitro experiment, two full—developed leaves from the top during heading stage were collected and cut into 5.0cm segments, They were floated on the surface of distilled water containing 0.1mg/14PU—30 and incubated in darkness at 30 C. The leaves floated on distilled water were used as control.It was observed that chlorophyll content in controlled leaves declined rapidly started from the second day and dropped by 93.4% on the 6th day while that in leaves treated with 4PU—30 declined by 41.4% only. During senescence, specific activities of hemoglobin—digesting

Effects of a GA3/4PU-30 compound (90-09) on leaf senescence of hybrid rice

90-09 is a cmpound made with GA3,4PU-30and some trace elements(B ete.).To studythe effects of 90-09 on the leaf senescence ofhybrid rice leaf,the changes of activities ofsome degradation enzyme and the contents ofendogenous hormones(ABA,ZRs,GAs,andIAA),protein,and nucleic acids were mea-sured during leaf senescence. Shanyou 63,an indica hybrid rice,wasused in this experiment.Rice plants were firstsprayed with 90-09(115 ml/hm~2) solution 10d after heading,followed by two more sprayswith once a week.The controlled plants weresprayed with water only.

Scaffold protein RACK1A positively regulates leaf senescence by coordinating the EIN3-miR164-ORE1 transcriptional cascade in Arabidopsis

Plants have adopted versatile scaffold proteins to facilitate the crosstalk between multiple signaling pathways. Leaf senescence is a well-programmed developmental stage that is coordinated by various external and internal signals. However,the functions of plant scaffold proteins in response to senescence signals are not well understood. Here, we report that the scaffold protein RACK1A(RECEPTOR FOR ACTIVATED C KINASE1A) participates in leaf senescence mediated by ethylene signaling via the coordination of the EIN3-miR164-ORE1 transcriptional regulatory cascade. RACK1A is a novel positive regulator of ethylene-mediated leaf senescence. The rack1a mutant exhibits delayed leaf senescence, while transgenic lines overexpressing RACK1A display early leaf senescence. Moreover, RACK1A promotes EIN3(ETHYLENE INSENSITIVE 3) protein accumulation, and directly interacts with EIN3 to enhance its DNA-binding activity. Together, they then associate with the miR164 promoter to inhibit its transcription, leading to the release of the inhibition on downstream ORE1(ORESARA 1) transcription and the promotion of leaf senescence.This study reveals a mechanistic framework by which RACK1A promotes leaf senescence via the EIN3-miR164-ORE1 transcriptional cascade, and provides a paradigm for how scaffold proteins finely tune phytohormone signaling to control plant development.

A molecular framework underlying low-nitrogeninduced early leaf senescence in Arabidopsis thaliana

Nitrogen(N)deficiency causes early leaf senescence,resulting in accelerated whole-plant maturation and severely reduced crop yield.However,the molecular mechanisms underlying N-deficiency-induced early leaf senescence remain unclear,even in the model species Arabidopsis thaliana.In this study,we identified Growth,Development and Splicing 1(GDS1),a previously reported transcription factor,as a new regulator of nitrate(NO3)signaling by a yeast-one-hybrid screen using a NO3enhancer fragment from the promoter of NRT2.1.We showed that GDS1 promotes NO3 signaling,absorption and assimilation by affecting the expression of multiple NO3 regulatory genes,including Nitrate Regulatory Gene2(NRG2).Interestingly,we observedthat gds1mutants show early leaf senescence as well as reduced NO3-contentand Nuptake under N-deficient conditions.Further analyses indicated that GDS1 binds to the promoters of several senescence-related genes,including Phytochrome-lnteracting Transcription Factors 4 and 5(PIF4 and PIF5)and represses their expression.Interestingly,we found that N deficiency decreases GDS1 protein accumulation,and GDS1 could interact with Anaphase Promoting Complex Subunit 10(APC10).Genetic and biochemical experiments demonstrated that Anaphase Promoting Complex or Cyclosome(APC/C)promotes the ubiquitination and degradation of GDS1 under N deficiency,resulting in loss of PIF4 and PiF5 repression and consequent early leaf senescence.Furthermore,we discovered that overexpression of GDS1 could delay leaf senescence and improve seed yield and N-use efficiency(NUE)in Arabidopsis.In summary,our study uncovers a molecular framework illustrating a new mechanism underlying low-N-induced early leaf senescence and provides potential targets for genetic improvement of crop varieties with increased yield and NUE.

ANAC087 transcription factor positively regulates age-dependent leaf senescence through modulating the expression of multiple target genes in Arabidopsis

Leaf senescence is the final stage of leaf development and appropriate onset and progression of leaf senescence are critical for reproductive success and fitness.Although great progress has been made in identifying key genes regulating leaf senescence and elucidating the underlining mechanisms in the model plant Arabidopsis,there is still a gap to understanding the complex regulatory network.In this study,we discovered that Arabidopsis ANAC087 transcription factor(TF)positively modulated leaf senescence.Expression of ANAC087 was induced in senescing leaves and the encoded protein acted as a transcriptional activator.Both constitutive and inducible overexpression lines of ANAC087 showed earlier senescence than control plants,whereas T-DNA insertion mutation and dominant repression of the ANAC087 delayed senescence rate.A quantitative reverse transcription-polymerase chain reaction(qRT-PCR)profiling showed that the expression of an array of senescence-associated genes was upregulated in inducible ANAC087 overexpression plants including BFN1,NYE1,CEP1,RbohD,SAG13,SAG15,and VPEs,which are involved in programmed cell death(PCD),chlorophyll degradation and reactive oxygen species(ROS)accumulation.In addition,electrophoretic mobility shift assay(EMSA)and chromatin immunoprecipitation-quantitative polymerase chain reaction(ChIP-qPCR)assays demonstrated that ANAC087 directly bound to the canonical NAC recognition sequence(NACRS)motif in promoters of its target genes.Moreover,mutation of two representative target genes,BFN1 or NYE1 alleviated the senescence rate of ANAC087-overexpression plants,suggesting their genetic regulatory relationship.Taken together,this study indicates that ANAC087serves as an important regulator linking PCD,ROS,and chlorophyll degradation to leaf senescence.

Melatonin retards leaf senescence by modulating phytohormone metabolism in stored Chinese flowering cabbage

This paper was conducted to explore the effects of melatonin(MT)on the senescence of stored Chinese flowering cabbage and the potential modulatory mechanisms involved.The physiological findings demonstrated that MT successfully reduced chlorophyll loss and improved the photochemical effectiveness of cabbage leaves.In addition,MT decreased the transcription of senescence-associated genes(BrSAG12)and genes responsible for chlorophyll breakdown.Transcriptome analysis showed that MT-regulated genes were enriched in oxidative phosphorylation,hormone metabolism and signal transduction,and MT treatment reduced the high expression of genes linked to generation of reactive oxygen species(ROS),energy metabolism,phytohormone(abscisic acid(ABA),ethylene(ET),and jasmonic acid(JA))biological synthesis and signal transduction while promoting the activation of genes related to scavenging ROS,energy biosynthesis and plant–pathogen interactions.We emphasized the examination of the potential interaction between phytohormone metabolism and MT.The results showed that the application of MT decreased ABA,ET,and JA levels as well as the expression of their biosynthesis genes,concurrently maintaining higher expression of cytokinin,auxin and gibberellin biosynthetic genes and lower expression of degradation genes.Regulatory networks of transcription factors(TFs)and genes related to ABA,ET and JA metabolism showed that TFs such as DNA-binding One Zinc Finger 5.7(DOF5.7),WRKY40,and homeobox-leucine zipper protein-16(ATHB-16)might play important transcriptional regulatory roles in mediating MT postponed leaf senescence.Taken together,these findings suggested that the postponed senescence of cabbage treated with MT might be ascribed to the regulated oxidative phosphorylation,energy,phytohormone metabolism,and transcription factors.

ABF2, ABF3, and ABF4 Promote ABA-Mediated Chlorophyll Degradation and Leaf Senescence by Transcriptional Activation of Chlorophyll Catabolic Genes and Senescence-Associated Genes in Arabidopsis

Chlorophyll （Chl） degradation is an integral process of leaf senescence, and NYE1/SGR1 has been demonstrated as a key regulator of Chl catabolism in diverse plant species. In this study, using yeast one-hybrid screening, we identified three abscisic acid （ABA）-responsive element （ABRE）-binding transcription factors, ABF2 （AREB1）, ABF3, and ABF4 （AREB2）, as the putative binding proteins of the NYE1 promoter. Through the transactivation analysis, electrophoretic mobility shift assay, and chromatin immunoprecipitation, we demonstrated that ABF2, ABF3, and ABF4 directly bound to and activated the NYE1 promoter in vitro and in vivo. ABA is a positive regulator of leaf senescence, and exogenously applied ABA can accelerate Chl degradation. The triple mutant of the ABFs, abf2abf3abf4, as well as two ABA-insensitive mutants, abil-1 and snrk2.2/2.3/2.6, exhibited stay-green phenotypes after ABA treatment, along with decreased induction of NYE1 and NYE2 expression. In contrast, overexpression of ABF4 accelerated Chl degradation upon ABA treatment. Interestingly, ABF2/3/4 could also activate the expression of two Chl catabolic enzyme genes, PAO and NYCl, by directly binding to their promoters. In addition, abf2abf3abf4 exhibited a functional stay-green phenotype, and senescence-associated genes （SAGs）, such as SAG29 （SWEET15）, might be directly regulated by the ABFs. Taken together, our results suggest that ABF2, ABF3, and ABF4 likely act as key regulators in mediating ABA-triggered Chl degradation and leaf senescence in general in Arabidopsis.

Gene Network Analysis and Functional Studies of Senescence-associated Genes Reveal Novel Regulators of Arabidopsis Leaf Senescence

Plant leaf senescence has been recognized as the last phase of plant development, a highly ordered process regulated by genes known as senescence associated genes （SAGs）. However, the function of most of SAGs in regulating leaf senescence as well as regulators of those functionally known SAGs are still unclear. We have previously developed a curated database of genes potentially associated with leaf senescence, the Leaf Senescence Database （LSD）. In this study, we built gene networks to identify common regulators of leaf senescence in Arabidopsis thaliana using promoting or delaying senescence genes in LSD. Our results demonstrated that plant hormones cytokinin, auxin, nitric oxide as well as small molecules, such as Ca2＋, delay leaf senescence. By contrast, ethylene, ABA, SA and JA as well as small molecules, such as oxygen, promote leaf senescence, altogether supporting the idea that phytohormones play a critical role in regulating leaf senescence. Functional analysis of candidate SAGs in LSD revealed that a WRKY transcription factor WRKY75 and a Cys2/His2-type transcription factor AZF2 are positive regulators of leaf senescence and loss-of-function of WRKY75 or AZF2 delayed leaf senescence. We also found that silencing of a protein phosphatase, AtMKP2, promoted early senescence. Collectively, LSD can serve as a comprehensive resource for systematic study of the molecular mechanism of leaf senescence as well as offer candidate genes for functional analyses.

Age-Triggered and Dark-Induced Leaf Senescence Require the bHLH Transcription Factors PIF3, 4, and 5

Leaf senescence can be triggered and promoted by a large number of developmental and environmental fac- tors. Numerous lines of evidence have suggested an involvement of phytochromes in the regulation of leaf senescence, but the related signaling pathways and physiological mechanisms are poorly understood. In this study, we initially identi- fied phytochrome-interacting factors （PIFs） 3, 4, and 5 as putative mediators of leaf senescence. Mutations of the PIF genes resulted in a significantly enhanced leaf longevity in age-triggered and dark-induced senescence, whereas overexpressions of these genes accelerated age-triggered and dark-induced senescence in Arabidopsis. Consistently, loss-of-function of PIF4 attenuated dark-induced transcriptional changes associated with chloroplast deterioration and reactive oxygen species （ROS） generation. ChlP-PCR and DualoLuciferase assays demonstrated that PIF4 can activate chlorophyll degradation regulatory gene NYE1 and repress chloroplast activity maintainer gene GLK2 by binding to their promoter regions. Finally, dark-induced ethylene biosynthesis and ethylene-induced senescence were both dampened in pif4, suggesting the involvement of PIF4 in both ethylene biosynthesis and signaling pathway. Our study provides evidence that PIF3, 4, and 5 are novel positive senes- cence mediators and gains an insight into the mechanism of light signaling involved in the regulation of leaf senescence.