The evolutionary basis of leaf senescence: method to the
madness?
Anthony B Bleecker
Recent studies on the differential expression of genes associated with leaf senescence support the long-standing interpretation of plant senescence as an organized, genetically controlled process. Sequence identities of genes that are differentially expressed in senescing leaves indicate roles in the salvage of nutrients. By considering this salvage function as the selected trait and the degeneration and death of the tissue a pleiotropic consequence of nutrient redistribution, the process of leaf senescence can be reconciled with evolutionary theories on the origins of senescence in animals.
Addresses
Botany Department, 430 Lincoln Drive, University of Wisconsin-Madison, Madison, WI 50706, USA; e-mail: Bleecker@facstaff.wisc.edu
Current Opinion in Plant Biology1998,1:73–78 http://biomednet.com/elecref/1369526600100073 Current Biology Ltd ISSN 1369-5266
Abbreviation
SAG senescence-associated gene
Introduction
Understanding the decline in vitality that characterizes the final stages in the life histories of most higher organisms may represent one of the greatest challenges to biological science and medicine. Yet, the field of senescence research struggles to define even the terms under which it should operate [1]. Is senescence the direct result of genetic programming, or is it the opposite — a general breakdown in systems that have passed the age where natural selective forces operate to fine tune them?
Much of the debate surrounding the nature of senescence is focused on animal models. The plant kingdom, however, provides its own set of unique properties and problems with respect to senescence. The apparent ability of some plant species to live indefinitely and the prevailing opinion that even somatic tissue senescence is a genetically pro-grammed process have set the study of plant senescence apart from the animal field [2••]. Have plants somehow escaped from the evolutionary constraints that have made limited lifespans a fact of life for most animals?
In this article, I will attempt to reconcile the apparent differences in how senescence in plants and animals is viewed from an evolutionary perspective. To this end, I will briefly summarize evolutionary theories that account for the prevalence of senescence in animals. I will then discuss some problems with comparing plant and animal life histories that arise from basic differences in the way
that plants and animals develop. Finally, I will suggest how the apparent programmed nature of leaf senescence can be reconciled with evolutionary theories of animal senescence by considering the individual leaf as the organizational unit on which selective forces have acted to create the senescence syndrome.
Senescence in animals and plants:
definitional dichotomy?
In order to consider the evolutionary origin of senescence in plants, some clarification of what is meant by the term senescence is necessary. This is a particular problem when discussing the relationship between senescence in plants and animals because the term senescence has been defined in different ways for these two classes of organisms. In an attempt to clarify this issue, I will first present a brief summary of the evolution-based definition of senescence that was developed from animal models but should apply to all higher organisms. I will then consider how the unique features of plants have led to some confusion in attempts to reconcile the evolutionary theories of senescence developed for animals with the developmental definition of senescence that is most often used to describe the process in plants.
pleiotropy was discussed more recently by Rose [3]. A different perspective was provided by the ‘disposable soma’ hypothesis [4], which posits that an organism’s maximum life span will reflect a tradeoff between the allocation of resources for the maintenance of somatic tissues of the adult and the preservation of the germline through the production of offspring. Given the greater importance of reproduction for survival of the genome, longevity of the individual is the inevitable loser.
The concepts of antagonistic pleiotropy and the disposable soma were developed from the study of animal systems where the soma is largely composed of differentiated, postmitotic tissues and where the germline is set aside early in development. Plants, however, present particular problems when attempting to apply the zoologically derived definition of senescence. The modular nature of plant growth and development and the propensity for veg-etative reproduction lead to difficulties in defining where an individual plant begins and ends. Even in the clearest cases of individual plants, the semiautonomous nature of plant organ systems presents difficulties in assigning a specific time of death for the whole organism. These attributes have caused scientists to make a distinction between a gamet, which represents all structures arising from a single seed, and a ramet, which represents some identifiable somatic structure as an individual (see [2••]). Individual shoot systems arising from an underground root system may be defined as ramets, and this definition has even been extended to the point where a single shoot may be thought of as a population of ramets composed of individual nodes [5]. From an evolutionary point of view it may be more appropriate to consider the life history of an individual leaf — a terminally differentiated, fairly autonomous entity — when attempting to reconcile the similarities and differences between plants and animals.
A second source of possible confusion with respect to plants is a semantic one. The term senescence has been used for many years in the plant literature to refer specifically to an orderly progression of genetically controlled events that lead to the degeneration and death of leaf tissues. Plant scientists have tended to emphasize the developmental aspects of leaf senescence, often using the term programmed senescence [2••,6••–8••]. The clear implication is that this programmed senescence is more analogous to programmed cell death or apoptosis than to the formal definition of senescence as a decline in function related to age. This interpretation is fueled by experimental evidence that the timing of leaf senescence can be influenced by hormonal and other developmental processes [2••,6••,7••] and by the long-standing evidence that gene expression is required for the senescence process (see [2••]). The recent that leaf senescence is developmentally regulated so permeates the literature that several authors have dismissed the notion that leaf senes-cence is related to the formal definition of evolutionary senescence as an age-related decline in function[1,2••,5].
This being the case, evolutionary theorists have had difficulty considering how such an active suicide program could have evolved in plants. Wilson [9••] considered several alternative explanations for the evolution of programmed senescence in plants, but was unable to develop a convincing hypothesis. On the other hand, my co-workers and I have suggested that leaf senescence in
Arabidopsis may be best explained using the concepts of antagonistic pleiotropy and the disposable soma [10].
The question of whether leaf senescence is related to the zoological definition of senescence or is strictly a genetically regulated progression of events is not just an academic one. The current emphasis on differential gene expression as a paradigm for the study of leaf senescence is predicated on the underlying assumption that senescence is a developmentally controlled phenomenon. If this assumption is essentially correct, then we can hope that molecular and genetic strategies will succeed in defining the underlying regulatory pathways that govern the process. If, on the other hand, leaf senescence is to some degree driven by the less well defined processes that are thought to contribute to the evolution of senescence in animals, then we might anticipate a more difficult time in sorting out the adaptive processes from chaotic events that are occurring because of the absence of selection pressure acting on the system.
Developmental aspects of the senescence
syndrome in leaves
Natural leaf senescence follows a predictable progression of events that are collectively referred to as the senescence syndrome (for recent reviews see [2••,11•]. Ultrastruc-tural changes are first noted in chloroplasts, where the breakdown of thylakoid membranes is associated with the visible loss of chlorophyll at the whole leaf level and a decrease in levels of chloroplast proteins at the biochemical level. In conjunction with the breakdown in chloroplast internal structure, decreases in cytoplasmic volume and in the number of cytoplasmic ribosomes are often noted. By contrast, the integrity of the plasma membrane, mitochondria and the nucleus is maintained until very late in the senescence process. The breakdown of chloroplast proteins and lipids is associated with a loss of carbon and nitrogen from the leaf as it senesces. The senescence syndrome, therefore, is interpreted in terms of nutrient salvage. Several lines of evidence indicate that this nutrient salvage system is a developmentally regulated program. Enucleation of the cell blocks chloroplast degen-eration, as do inhibitors of protein and RNA synthesis (see [2••,6••]). Chloroplast degeneration is also reversible in some experimental systems [2••]. These observations have led to the recent interest in applying the techniques of molecular biology to define the role that differential gene expression may be playing in the senescence syndrome.
hybridization to identify cDNA clones that represent genes that are differentially expressed during senescence. Recent reviews provide detailed surveys of sequenced genes and their expression patterns [2••,6••–8••,11•], so I will only provide a brief summary here. Three broad classes of cDNAs can be seen based on expression patterns: first, those that decrease in relative abundance; second, those that show little change in relative abun-dance; and third, those that increase in relative abundance as senescence progresses. A caveat to all of these studies is the use of total RNA to determine relative mRNA abundance which can be misleading when one considers that total RNA declines significantly during senescence. This problem has been addressed in some studies by calculating mRNA abundance on a per leaf basis [10,12].
Many of the genes classified as senescence-associated genes (SAGs) are expressed in green tissues and increase in relative abundance at later stages of senescence. Some of these may simply be genes whose levels of expression are maintained in cells while levels of most mRNA classes and rRNAs are declining. A smaller set of cloned cDNAs represents genes that are more clearly expressed only in senescing cells [7••,12]. The distinction between these two sets of genes may be important in that although the former may increase in relative abundance during senescence (as a consequence of message stability), the latter are better candidates as genes that are transcrip-tionally activated during senescence. Attempts to study expression patterns of SAGs in detail led in one case to the classification of nine different patterns of expression [7••]. This level of complexity may reflect the operation of many different pathways operating by different mechanisms.
Sequence analysis of SAGs has resulted in the iden-tification of a number of genes that are likely to be involved in salvage functions [2••,6••–8••,11•]. These include genes encoding proteases, nucleases, lipid- , carbohydrate- and nitrogen-metabolizing enzymes and other enzymes involved in mobilization of nutrients. Another class of sequences appears to code for protective or stress response functions [7••]. The latter may act to protect the increasingly fragile cell while the task of salvage is being completed, or they may simply respond blindly to the trauma caused by salvage and perform no adaptive function at all. Finally, many cloned SAG sequences have no obvious role in salvage or protection [7••,11•]. Is the expression of these genes significant or simply the molecular ramblings of cells going senile?
One class of genes that are conspicuously absent from SAG populations is those with obvious gene-regulatory functions. For example, no transcription factor candidates have been reported. It is possible that messages for differentially expressed transcription factors are present but too rare to have been picked up by current techniques.
Alternatively, if specific transcription factors are required at all, they may be present in nonsenescing cells and activated by whatever decision-making machinery must by definition be present in presenescent tissues. It is even conceivable that decreased expression of transcriptional repressors is the key regulatory event. In any case, sequence analysis of some SAG 5′regulatory regions has not revealed any common sequence elements between different SAGs [6••,7••]. Taken together with the wide range of expression patterns observed for SAGs, these results support the idea that the proximal regulation of gene expression associated with senescence may be complex [6••,8••,9••]. This may also be why mutational analysis inArabidopsishas yet to provide any mutants that block the senescence process [13••].
Lacking a clearly developed picture for proximal regula-tory systems, I turn now to the consideration of higher order regulatory systems that appear to control the timing and progression of leaf senescence. Reminiscent of the proverbial blind men and the elephant, one’s concept of how leaf senescence is controlled depends very much on which plant system one is examining. The relevant systems have been the subjects of recent reviews so I will just mention them briefly here. There are clearly examples where specific hormones have a controlling influence over organ senescence. Cytokinins can have a dramatic effect on delaying leaf senescence in some species [14]. The effectiveness of applied cytokinin, however, varies considerably between plant species. Ethylene promotes senescence in many species but several recent studies indicate that ethylene is neither necessary nor sufficient to cause senescence in Arabidopsis or tomato [15,16•]. On the other hand ethylene is the key regulatory factor in the senescence of floral organs in some species [17]. Environmental signals can also trigger senescence. Shad-ing of lower leaves by upper leaves induces senescence in some species (see [6••]), while deciduous trees are affected by temperature and day length. Reproductive development can trigger senescence of vegetative tissue both proximally and even on a global scale in monocarpic species. Evidence from soybean is consistent with a chemical communication system between fruits and leaves that triggers the senescence syndrome [2••]. Finally, leaf age may be the best predictor of when senescence will occur; this seems to be the case inArabidopsis[10].
Reconciling the evolutionary origin of
senescence in plants and animals
In considering the evolutionary origin of leaf senes-cence, I propose that the senescence syndrome, or more specifically the salvage system that it represents, is a primitive set of pathways that were recruited early in the evolution of land plants because these biochemical pathways contributed to the retention of nutrients by the whole plant. I use the term recruited because it seems plausible that the biochemical activities required for the operation of the senescence syndrome were already functioning at reduced levels as a component of cellular maintenance. This may explain why so many SAGs are still expressed at lower levels in healthy tissue. Perhaps the senescence syndrome evolved by mechanisms that shifted the balance of anabolic and catabolic components of cell metabolism in favor of catabolism. A key element of this proposal is the idea that the senescence syndrome was originally selected to salvage nutrients, not to cause the death of the tissue in which it is expressed.
It is my contention that the many facets of the senescence syndrome discussed above can be brought into perspective and reconciled with the evolutionary theories of senes-cence developed for animal systems if one considers the individual leaf as the organizational unit on which natural selection acted thus creating the senescence syndrome. Obviously, the somatic tissues of the leaf do not contribute directly to the future of the genetic line, but then, neither do the somatic tissues of an animal. Leaves do provide an indirect contribution to the genetic line in the form of photosynthetic assimilates. One would expect natural selection to favor the continued survival of a leaf as long as this assimilatory function is operable.
Despite this expectation a variety of environmental and developmental factors may limit the functional ‘life expectancy’ of a leaf. Predation, sporadic and seasonal climate extremes, and competition for light — both from competing plants and ultimately from continued growth of the parent shoot — may contribute to a diminishing capacity of a leaf to maintain photosynthetic output. According to the disposable soma hypothesis [4] it is unlikely that natural selection would favor maintaining leaves in a healthy state when they age beyond the functional life expectancy imposed by these external factors. One would expect that natural selection would favor the evolution of salvage pathways that are either activated or enhanced as leaves approach this maximum functional life expectancy because these salvage systems would produce a continued contribution to the plant in the form of salvaged nutrients.
The massive nutrient salvage associated with the senes-cence syndrome and cell maintenance are not compatible programs. A pleiotropic consequence of the salvage pathways is that their activities tend to increase the mortality rate of the tissues in which they are expressed.
Natural selection will not have a correcting influence on these negative consequences of salvage if the leaves at that stage have a low probability of contributing to reproduction via photosynthesis. In other words, salvage pathway genes fit the theory of antagonistic pleiotropy [1,4]. They were selected during evolution for their contributions to reproductive success, but have a negative impact on the longevity of the somatic tissues in which they operate.
Cell death: developmental program or
gratuitous pogrom?
If one accepts the idea that the primal function of the senescence syndrome was salvage, then the subsequent death of the leaf could be considered a consequence of the lack of selection for continued survival. From this point of view, the senescence syndrome cannot be considered a cell death program, although this extreme theoretical viewpoint only considers positive contributions of the leaf to the whole plant. A leaf that is left in a weakened state as a result of salvage could become a liability to the whole plant, for example, as a target for pathogen invasion. In this case, evolutionary forces could select for pathways that deliver the coup de grace to senile leaves. Such pathways could be selected by recruitment of specific subroutines within the overall syndrome. They could also evolve as completely different developmental programs, the abscission process being the primary example [10].
The plant leaf is a semiautonomous unit. It receives information from its own internal processes and directly from the environment. The leaf also receives chemical signals from the parent plant that carry information as to the nutritional and developmental status of the plant. Information processing networks that have evolved within the leaf could couple any combination of these signals to cause the activation of salvage pathways. The developmental control of these salvage pathways opens the possibility that they could be co-opted by evolution to function more directly in programmed cell death. For example, the rapid senescence of floral displays after fertilization [18] may have been selected more as a protection against predators than as a mechanism of salvage. Even the monocarpic senescence of crop plants like soybean and pea [19] could be the result of selective breeding for ease of human harvest; in which case the target of selection would be the global death of vegetative tissues rather than salvage.
Conclusion
syndrome is explained if one assumes that the adaptive significance of the syndrome is in nutrient salvage and that the associated cell death is a pleiotropic consequence of the program. Once these salvage pathways are in place, however, they could be co-opted by natural selection to function more directly in the death of tissues and organ systems.
The study of gene expression associated with senescence indicates that the process is multigenic and complex. No clear picture of the genetic regulatory mechanisms that that drive the process has yet emerged out of these studies. The overall syndrome may best be viewed as a collection of parallel and branched regulatory pathways that may form a regulatory grid. These salvage pathways may be activated when the leaf no longer contributes sufficiently to photosynthetic function, or they may be coupled to de-velopmental signals that activate programmed senescence in otherwise healthy tissues.
Challenges facing the study of leaf senescence involve characterizing the various pathways and the specific functions they regulate. The genetic approach provides the greatest promise to produce definitive answers in this regard. Mutations in SAG genes and in the pathways that regulate them will help to sort out the different regulatory pathways involved. Various approaches for achieving this goal using Arabidopsis as a model system have been discussed [6••,10]. Specific examples where genetics has provided interesting insights include the stay-green mutants inFestuca[18], which are blocked in the turnover of chlorophyll in the thylakoid membranes but otherwise show most aspects of the senescence syndrome. In soybean, mutations at two loci, designated d1 and d2, cause a delay in leaf senescence. A maternally inherited mutation, cytG, has a similar effect. The combination of these mutants appears to block the ability of reproductive structures to trigger the senescence syndrome in other-wise healthy soybean leaves [2••,19]. Interestingly, these mutations do not block the activation of abscission in these leaves. These examples demonstrate that mutations which affect specific subroutines or signalling pathways associated with the overall senescence syndrome can help to unravel the complexities of senescence in plants.
References and recommended reading
Papers of particular interest, published within the annual period of review, have been highlighted as:
• of special interest •• of outstanding interest
1. Finch CE:Longevity, Senescence, and the Genome.Chicago: University of Chicago Press: 1990.
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2. Nood ´en LD, Guiamet JJ:Genetic control of senescence and aging in plants. InHandbook of the Biology of Aging. Fourth edition. Edited by Schneider EL, Rowe JW. San Diego: Academic Press; 1996:94-118.
The authors provide a summary of the field of plant senescence from several perspectives and include a survey of genes that are differentially expressed during senescence. They argue that, because leaves may senesce before or during reproduction, evolutionary models for animal senescence do not
apply. This may seem true from a strictly whole-plant perspective, but not when one considers the individual leaf as a unit of selection.
3. Rose MR:Evolutionary Biology of Aging. Oxford: Oxford University Press; 1991.
4. Kirkwood TB, Rose MR:Evolution of senescence: late survival sacrificed for reproduction.Philos Trans R Soc Lond1991,
332:15-24.
The authors provide an interesting and readable discussion of two prominent theories that account for the evolution of senescence in animals: antagonistic pleiotropy and the disposable soma.
5. Roach DA:Evolutionary senescence in plants.Genetica1993,
91:53-64. ••
6. Gan S, Amasino RM:Making sense of senescence.Plant Physiol
1997,113:313-319.
This recent review of leaf senescence both considers the recent studies on gene expression and summarizes the developmental and environmental pathways that influence the timing of senescence in different systems. The senescence syndrome is presented as a form of programmed cell death, but the authors do consider the apparent complexity of regulatory factors that may operate in the syndrome and suggest a network of parallel and overlapping pathways that control the process (Figure 3).
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7. Buchanan-Wollaston V:The molecular biology of leaf senescence.J Exp Biol1997,307:181-199.
The author focuses on the analysis of differential gene expression during leaf senescence. This is perhaps the most thorough of recent reviews on this topic. A flow chart of salvage pathways thought to be active during leaf senescence is provided and reconciled with the identities of many senescence-associated gene clones. An attempt is made to classify genes according to their expression patterns, of which 10 different classes are identified. It is recognized that the complexity of expression patterns may indicate a plethora of regulatory pathways. Yet, the author is convinced that transcriptional control is the key to senescence and that all genes which are differentially regulated must be important to the process.
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8. Nam HG:The molecular genetic analysis of leaf senescence. Curr Opin Biotechnol1997,8:200-207.
In the same vein as other recent reviews [7••], the author use gene expres-sion as the paradigm of control. An emphasis is placed here on the idea that hormonal and developmental pathways overlap in controlling different senescence-associated genes (SAGs) and that different forms of senes-cence (starvation induced, hormonally induced and age related) show dif-ferences in which SAGs are elevated. In the conclusion, the importance of considering post-transcriptional mechanisms for SAG abundance is empha-sized. Examples where mRNA and protein abundance data do not neces-sarily agree are also noted as a cautionary point.
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9. Wilson BJ:An evolutionary perspective on the ‘death hormone’ hypothesis in plants.Physiol Plant1997,99:511-516.
The author considers the problem of how a developmental system that causes death could have evolved. Selection at several different levels is con-sidered and the author concludes that nutrient competition between leaves and fruits is the best candidate. Wilson discounts this mechanism, however, because even old leaves are not net importers so their death is of no conse-quence; and also, monocarpic varieties do not show a consistent advantage over their polycarpic relatives in terms of yield. To the first argument, I would respond that export from older leaves, not competition for import, is the issue. Release of stored nitrogen and other nutrients from otherwise unproductive leaves is the trait under consideration. The second argument, therefore, is based on flawed experiments. Comparing varieties with multiple genetic dif-ferences in species that have been artificially selected by humans and grown under specialized protected conditions is not likely to be the criterion that evolutionary processes would use in selecting the trait.
10. Hensel LL, Grbic V, Baumgarten DA, Bleecker AB:Developmental and age related processes that influence the longevity and senescence of photosynthetic tissues in Arabidopsis.Plant Cell
1993,5:553-564. •
11. Weaver LM, Himelblau E and Amasino RM:Leaf senescence: gene expression and regulation.InGenetic Engineering, V.19: Principles and Methods.New York: Plenum; in press.
This is a thorough review of leaf senescence that considers primarily the biochemical and molecular aspects of the process.
12. Lohman KN, Gan S, John MC, Amasino RM:Molecular analysis of natural leaf senescence inArabidopsis thaliana.Physiol Plant1994,92:322-328.
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13. Bleecker AB, Patterson SE:Last exit: senescence, abscission and meristem arrest inArabidopsis.Plant Cell1997,9 :1169-1179.
plant senescence. Theoretical discussions relevant to this paper are also included.
14. Gan S and Amasino RM:Inhibition of leaf senescence by auto-regulated production of cytokinin.Science1995,270 :1986-1988.
15. John I, Drake R, Farrell A, Cooper W, Lee P, Horton P, Grierson D:Delayed leaf senescence in ethylene deficient ACC-oxidase antisense tomato plants. Molecular and physiological aspects. Plant J1995,7:483-490.
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16. Grbic V, Bleecker AB:Ethylene regulates the timing of leaf senescence inArabidopsis.Plant J1996,8:595-602.
The authors use ethylene insensitive mutants to investigate the role of ethy-lene in the timing of leaf senescence. Delayed leaf senescence was observed
but when it did occur it was associated with wild-type levels of senescence-associated gene mRNAs. It was concluded that ethylene was neither nec-essary nor sufficient to cause leaf senescence, and that it only influenced the timing.
17. O’Neill S, Nadeau JA, Zhang XS, Bui AQ, Halevey AH: Inter-organ regulation of ethylene biosynthetic genes by pollination. Plant Cell1993,5:419-432.
18. Thomas H, Smart CM:Crops that stay green.Ann Appl Biol
1993,123:193-219.
