Hormonal regulation of leaf senescence through integration of developmental and stress signals

Leaf senescence is a genetically controlled dismantling programme that enables plants to efficiently remobilise nutrients to new growing sinks. It involves substantial metabolic reprogramming whose timing is affected by developmental and environmental signals. Plant hormones have long been known to affect the timing of leaf senescence, but they also affect plant development and stress responses. It has therefore been difficult to tease apart how the different hormones regulate the onset and progression of leaf senescence, i.e., whether they directly affect leaf senescence or affect it indirectly by altering the developmental programme or by altering plants’ response to stress. Here we review research on hormonal regulation of leaf senescence and propose that hormones affect senescence through differential responses to developmental and environmental signals. We suggest that leaf senescence strictly depends on developmental changes, after which senescence can be induced, depending on the type of hormonal and environmental cues.     

Molecular regulation of leaf senescence

Leaf senescence is a process of programmed cell death, which is induced in an age-dependent manner and by various environmental cues. The mechanisms that regulate the induction and progression of leaf senescence remain unclear because of their complexity. However, recent genetic and reverse-genetic approaches have identified key components of the regulation of leaf senescence and have revealed glimpses of the underlying molecular mechanisms.     

Hormonal regulation of leaf senescence in Lilium

In addition to floral senescence and longevity, the control of leaf senescence is a major factor determining the quality of several cut flowers, including Lilium, in the commercial market. To better understand the physiological process underlying leaf senescence in this species, we evaluated: (i) endogenous variation in the levels of phytohormones during leaf senescence, (ii) the effects of leaf darkening in senescence and associated changes in phytohormones, and (iii) the effects of spray applications of abscisic acid (ABA) and pyrabactin on leaf senescence. Results showed that while gibberellin 4 (GA(4)) and salicylic acid (SA) contents decreased, that of ABA increased during the progression of leaf senescence. However, dark-induced senescence increased ABA levels, but did not affect GA(4) and SA levels, which appeared to correlate more with changes in air temperature and/or photoperiod than with the induction of leaf senescence. Furthermore, spray applications of pyrabactin delayed the progression of leaf senescence in cut flowers. Thus, we conclude that (i) ABA plays a major role in the regulation of leaf senescence in Lilium, (ii) darkness promotes leaf senescence and increases ABA levels, and (iii) exogenous applications of pyrabactin inhibit leaf senescence in Lilium, therefore suggesting that it acts as an antagonist of ABA in senescing leaves of cut lily flowers.     

The interaction of hormonal and environmental factors in leaf senescence

The work concerns the senescence of isolated young leaves of oats (Avena sativa) floated on water or solutions. Senescence is rapid in darkness but slow in white light; the effect of light is not due to photosynthesis, but is paralleled by stomatal opening. Closure of the stomata by osmotic or chemical means makes senescence in light proceed as fast as in darkness, while opening the stomata in darkness by cytokinins, fusicoccin,etc., delays senescence to rates typical of light. The osmotic closure in light is mediated by abscisic acid, and since this also accumulates in darkness it appears as a major factor controlling senescence. Efflux of ions into the solution; indicating increased permeability, occurs almost in parallel with senescence. Senescence in light is accelerated by 1-aminocyclopropane-l-carboxylic acid (ACC) and inhibited by cobalt, silver or aminoethoxyvinyl glycine (AVG) which interfere with ethylene production or action; however, ethylene’s role is unclear because some reagents, including kinetin, that delay senescence, actually increase ethylene production. At the endogenous level, therefore, ethylene may not be a limiting factor. Finally, a new ethylene-generating system is described in which the dehydrogenation of linoleic acid is coupled through manganese to the oxidation of ACC; it is probably activein vivo.     

The role of the epidermis in kinetin-induced retardation of chlorophyll degradation in tobacco leaf discs during senescence

Three kinds of discs were taken from tobacco leaves whose lowerepidermis had been peeled off, half-peeled or unpeeled. Therole of the epidermis and its relation to the kinetin effecton chlorophyll degradation during senescence were studied. Ourresults follow.

1. Chlorophyll degradation due to kinetin was retarded only whenthe lower epidermis was present.
2. The decrease in chlorophyllcontent in leaf discs on water duringsenescence was nearlyproportional to the size of the lowerepidermis attached tothe discs; i.e., unpeeled discs>half-peeleddiscs>peeleddiscs.
3. Cellular fractions possessing activity which induceschlorophylldegradation were extracted from the isolated lowerepidermis(i, ii) and its acetone powder (iii): (i) L-2 fraction(1.14d1.16)was separated by stepwise sucrose density-gradientcentrifugationfrom the 10,000?g pellet of the cell homogenate.(ii) The A-fraction(M.W.5,000) was precipitated with 0–80%saturation ofammonium sulfate from 105,000 ? g supernatantof cell homogenateand eluted in the void volume by SephadexG-25 column chromatography.(iii) The fraction precipitatedwith 0–30% saturationof ammonium sulfate from the 105,000?gsupernatant, containeda large amount of DNA and its activityremained even if DNAwas removed.
4. Activity was not retainedwhen the fractions were obtained fromisolated lower epidermispretreated with 2?10^–5 M kinetinfor 2 hr in darknessat 25?C.

(Received June 3, 1976; )     

The conserved mobility of mitochondria during leaf senescence reflects differential regulation of the cytoskeletal components in Arabidopsis thaliana

Leaf senescence is an organized process, which requires fine tuning between nuclear gene expression, activity of proteases and the maintenance of primary metabolism. Recently, we reported that leaf senescence was accompanied by an early degradation of the microtubule cytoskeleton in Arabidopsis thaliana. As the cytoskeleton is essential for cell stability, vesicle shuttling and organelle mobility, it might be asked how the regulation of these cell functions occurs with such drastic modifications of the cytoskeleton. Based on confocal laser microscopy observations and a micro-array analysis, the following addendum shows that mitochondrial mobility is conserved until the late stages of leaf senescence and provides evidences that the actin-cytoskeleton is maintained longer than the microtubule network. This conservation of actin-filaments is discussed with regards to energy metabolism as well as calcium signaling during programmed cell death.Key words: actin, cytoskeleton, microtubule, mitochondria, mobility, senescence     

Protease activity during rice leaf senescence

A protease activity was detected in rice (Oryza sativa L. cv. Ratna) leaves that hydrolysed hemoglobin more efficiently than bovine serum albumin. The activity was high when the enzyme was extracted and assayed with tris-maleate buffer [tris (hydroxymethyl) methyl amino-maleate] pH 7.0 rather than with water or with citrate-phosphate buffer pH 7.0. The enzyme had a strong dependence on sulfhydryl groups for its activity without which it was inaotive. The pH optimum was 7.0 and the temperature optimum was 40 °C. Protease activity expressed per unit leaf fresh weight (absolute activity) increased only little during senescence of detached rice leaves while the same activity expressed per unit soluble protein content (specific activity) increased by a greater factor (about 5 times) than absolute activity. Total and soluble protein content decreased during the senescence of detached leaves. Benzimidazole (10-3M) and kinetin (0.5xl0-5M) treatment arrested the increase of the protease activity and the deorease in the protein content during detached leaf senescence. It was indicative that protease protein was more stable than the bulk of other proteins in senescing leaves.     

Interactions of abscisic acid and sugar signalling in the regulation of leaf senescence

Leaf senescence can be triggered by a high availability of carbon relative to nitrogen or by external application of abscisic acid (ABA). Most Arabidopsis mutants with decreased sugar sensitivity during early plant development are either ABA insensitive (abi mutants) or ABA deficient (aba mutants). To analyse the interactions of carbon, nitrogen and ABA in the regulation of senescence, wild-type Arabidopsis thaliana (L.) Heynh. and aba and abi mutants were grown on medium with varied glucose and nitrogen supply. On medium containing glucose in combination with low, but not in combination with high nitrogen supply, senescence was accelerated and sucrose, glucose and fructose accumulated strongly. In abi mutants that are not affected in sugar responses during early development (abi1-1 and abi2-1), we observed no difference in the sugar-dependent regulation of senescence compared to wild-type plants. Similarly, senescence was not affected in the sugar-insensitive abi4-1 mutant. In contrast, the abi5-1 mutant did exhibit a delay in senescence compared to its wild type. As ABA has been reported to induce senescence and ABA deficiency results in sugar insensitivity during early development, we expected senescence to be delayed in aba mutants. However, the aba1-1 and aba2-1 mutants showed accelerated senescence compared to their wild types on glucose-containing medium. Our results show that, in contrast to sugar signalling in seedlings, ABA is not required for the sugar-dependent induction of leaf senescence. Instead, increased sensitivity to osmotic stress could have triggered early senescence in the aba mutants.Abbreviations ABA Abscisic acid - aba Abscisic acid deficient - abi Abscisic acid insensitive - F_v/F_m Maximum efficiency of photosystem II photochemistry     

Activation of latent phenolase during spinach leaf senescence

The observed increase of phenolase activity and of its rate of activation during spinach leaf senescence is due to reduced binding of latent phenolase to the thylakoid membranes and not to de novo synthesis. The same amount of phenolase which is active in isolated thylakoid membranes from senescent leaves can be found in the membranes of non-senescent leaves after activation of latent enzyme. Tracer experiments give evidence that one multiple form which is responsible for the bulk activity in senescent leaves, is synthesized before, but not after the onset of senescence, indicating that pre-existing latent phenolase is converted to easily activating forms.     

The molecular biology of leaf senescence

Senescence is a complex, highly regulated, developmental phasein the life of a leaf that results in the co-ordinated degradationof macromolecules and the subsequent mobilization of componentsto other parts of the plant. The application of molecular biologytechniques to the study of leaf senescence has, in the lastfew years, enabled the isolation and characterization of a largerange of cDNA clones representing genes that show increasedexpression in senescing leaves. The analysis of these genesand identification of the function of the encoded proteins willallow a picture of the complex processes that take place duringsenescence to be assembled. To date, genes encoding degradativeenzymes such as proteases and nucleases, enzymes involved inlipid and carbohydrate metabolism and enzymes involved in nitrogenmobilization have all been identified as senescence-enhancedgenes. A variety of other genes of no obvious senescence-relatedfunction have also been identified; their role in senescencemay be less predictable and, possibly, more interesting. The combined action of several internal and external signalsmay be involved in the induction of senescence. Analysis ofthe regulatory mechanisms controlling the expression of senescence-inducedgenes will allow the signalling pathways that are involved inthe regulation of senescence to be elucidated. Experiments withtransgenic plants and mutants are already shedding light onthe role played by cytokinins and ethylene in regulating senescencein leaves. Key words: Senescence, cDNA clones, gene expression, signals