Senescence in Plants Introductory article

Vicky Buchanan-Wollaston, University of Warwick, Wellesbourne, UK Article Contents

. Introduction
Senescence is the final stage of plant development during which the plant reclaims the . Changes that Occur during Senescence
valuable cellular building blocks that have been deposited in the leaves and other parts of . Gene Expression during Senescence
the plant during growth. Maintaining an efficient senescence process is essential for . Regulation of Leaf Senescence
survival of the plant or its future generations. Senescence is a complex highly regulated . Value of Senescence Research
process that requires new gene expression and involves the interactions of many signalling
pathways. doi: 10.1002/9780470015902.a0020133

Introduction
Senescence in plants is a complex, developmentally regu-
Changes that Occur during Senescence
lated phase during which cellular structures are carefully
Once a leaf is destined for senescence, it enters a highly
dismantled and the soluble components transferred to
regulated programmed series of events by which its cellular
other parts of the plant, either for storage or to promote
components are dismantled, degraded and mobilized. In
further growth. All parts of the plant undergo senescence
the poplar analysis described above, it was shown that over
but for the purpose of this article the emphasis will mainly
80% of the nitrogen and phosphorous were reclaimed from
be on the senescence of leaves since this has been the most
the senescing leaves. This process is controlled and usually
studied in recent years.
the leaf is maintained in a viable state until the remobili-
Leaf senescence occurs when the leaf is no longer of
zation is complete. The high degree of control is shown
use to the plant. This may be due to the stage of develop-
clearly by the fact that senescence can be reversed, if the
ment of the plant, the age of the leaf or be induced by
process has not proceeded beyond a certain stage.
environmental factors. As plants enter the flowering
Chloroplasts can be induced to regreen and to start pho-
and seed development stage, the nutrients within the leaves
tosynthesizing again.
are required for the seed development and so, in this
Loss of chlorophyll, with the resulting yellowing of the
case, senescence is induced by developmental signals.
leaves is the most obvious sign of senescence and in some
Alternatively, plant growth results in the leaves at the base
cases, as seen in Figure 1a and c, the additional synthesis
of the plant becoming shaded and restricted in their pho-
of anthocyanins during senescence results in dramatic
tosynthetic potential, in this case senescence is induced by
orange, red and even purple coloration. Synthesis of
environmentally controlled signals. Other types of envi-
anthocyanins by certain species has been shown to be
ronmental stress such as water and nutrient limitation or
important to protect the leaf from photooxidative damage
pathogen attack can also affect plant growth and bring
and maintain viability as senescence progresses. Another
about premature senescence. Senescence is at its most
noticeable fact of leaf senescence is illustrated in the leaves
obvious in the autumn when the dramatic colours of decid-
shown in Figure 1c. Often the leaf does not senesce evenly;
uous trees are striking reminder of the changing seasons
the tip and edges usually show the first signs with the areas
(Figure 1a). This senescence is caused by seasonal changes
around the vein tissues and the end of the leaf nearest the
such as light and temperature. A detailed molecular study
plant being the last to senesce. The reason for this is
in poplar in Sweden has indicated that senescence starts on
obvious if it is considered that the purpose of senescence is
exactly the same day each year irrespective of environ-
to remove soluble components from the leaf, via the vein
mental condition and follows the same degradative path-
system, before the leaf is allowed to die and drop off the
ways. This implies that, for this species, photoperiod is the
plant. Cells around the veins are kept active until maxi-
sole regulator causing the onset of senescence.
mum mobilization has occurred.
In perennial plants and trees, the mobilized components
are stored in roots or trunks to provide the building blocks
for growth in the following year. In annual plants, such as Structural changes
wheat, soybean and the model plant species, Arabidopsis,
the entire plant undergoes senescence and all reusable The majority of the protein and lipid component of a leaf is
nutrients are stored in the seed (Figure 1b). in the chloroplast and this is where the first and most

ENCYCLOPEDIA OF LIFE SCIENCES & 2007, John Wiley & Sons, Ltd. www.els.net 1
 Senescence in Plants

Figure 1 Illustrations of plant senescence: (a) Autumnal senescence in a beech wood, Derbyshire. (b) Total senescence in the entire wheat plant results in
all mobilizable nutrients being stored in the grain. (c) Leaf senescence showing differential progression of senescence in the leaf. Areas close to the veins
senesce last.

obvious structural changes occur during senescence. The Chlorophyll degradation

highly organized thylakoid membrane stacks, characteris-
tic of chloroplasts in mature green leaves become dis- Leaf yellowing is the first visible sign of senescence but chlo-
persed and by mid-senescence, the membrane structure rophyll levels have started to drop well before yellowing is
has become totally disorganized. The formation of large clearly visible. Figure 2 shows the changes in chlorophyll lev-
electron dense globules called plastoglobuli is characteris- els in the same Arabidopsis leaf harvested at different time
tic of the senescence of chloroplasts. These bodies are points during development and it is clear that 50% of chlo-
thought to have a role in lipid mobilization but may also rophyll is lost before there are obvious signs of yellowing in
be involved in the degradation of chlorophyll. Mito- the leaf. In mature green leaf cells, chlorophyll is bound to
chondria provide essential energy to maintain the protein apoproteins in the thylakoid and this complex is responsible
synthesis and enzyme activity required for senescence for light trapping during photosynthesis. During senescence,
to progress and these organelles are maintained until the the N content of these apoproteins (which comprises around
final stages. As senescence progresses, the plasma and vac- 30% of the total plastid nitrogen) must be remobilized, and
uolar membranes finally become disrupted and cell death chlorophyll is released. The first steps of chlorophyll deg-
occurs. radation are common to all plants and involve several

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 Senescence in Plants

1.2

Chlorophyll A+B
0.8

0.6

0.4

0.2

0
Day 20 Day 22 Day 24 Day 26 Day 28 Day 30 Day 32 Day 34 Day 36 Day 38 Day 40 Day 42
(a) Days after sowing

Day 20 Day 22 Day 24 Day 26 Day 28 Day 30 Day 32 Day 34 Day 36 Day 38 Day 40 Day 42
(b) Days after sowing

10

0
Day 20 Day 22 Day 24 Day 26 Day 28 Day 30 Day 32 Day 34 Day 36 Day 38 Day 40
(c) Days after sowing

Figure 2 Changes in the Arabidopsis leaf during development. (a) Change in chlorophyll levels as senescence progresses (b) Change in visual appearance
of leaf 7 harvested from Arabidopsis Col0 plants at different time points during development (c) Gene expression changes during leaf development.
GeneSpring (Silicon Genetics) analysis shows two clusters of genes each line shows the changes in expression of a single gene, those in red show increased
expression during development while those in blue show a decrease. SAG12 is one of the genes in the red cluster, several chlorophyll binding proteins and
other photosynthetic proteins are in the blue cluster.

enzymatic steps leading to the production of colourless chlorophyll catabolite reductase (RCCR) has been an essen-
compounds (primary fluorescent chlorophyll catabolites, tial step in the elucidation of the degradation pathway.
pFCCs). Following this the processes are different in differ- Once chlorophyll is separated from the proteins that
ent species, pFCCs undergo various modifications before bind it in the chloroplast membranes it becomes highly
being transported from the chloroplast and stored in the toxic, being very reactive to light. Therefore it is very
vacuole as nonfluorescent chlorophyll catabolites (NCCs). important that chlorophyll is rapidly degraded as a pro-
Chlorophyll degradation starts with the disruption of the tective measure against phototoxicity. Interestingly, two of
thylakoid structure allowing access to the proteins that bind the key enzymes in the chlorophyll degradation pathway
chlorophyll in the membrane. Chlorophyllase catalyses the (POA and RCCR) were identified originally in Arabidopsis
conversion of chlorophyll into chlorophyllide which is then mutants showing a light-dependent cell-death phenotype.
converted into pheophorbide a, followed by red chlorophyll In the absence of the complete chlorophyll-degradation
catabolite (RCC) and then to the pFCC intermediate. The pathway, the build up of toxic intermediates results in cell
genes involved in the pathway for chlorophyll degradation death and plants lacking these enzymes are extremely vul-
in plants have been identified in the last few years. The nerable to light stress. Therefore having a functional chlo-
cloning of the genes encoding the three key enzymes rophyll degradation pathway is vitally important for plant
chlorophyllase, pheophorbide aoxygenase (PAO) and red development and survival.

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 Senescence in Plants

Protein degradation hundreds of senescence-enhanced genes have been identified

and these are implicated as encoding proteins that have a
Proteins are degraded during leaf senescence to release val- role in the senescence process. However, the actual function
uable nitrogen and other minerals for mobilization. The level for the vast majority of these genes remains a mystery. Mi-
of total protein falls rapidly during leaf senescence, and this is croarray experiments using Arabidopsis gene probes and
paralleled by an increase in transcription and activity of a RNA isolated from green and senescent Arabidopsis leaves
number of different types of protease including vacuolar have shown that at least 800 genes are increased in expres-
processing enzymes. The majority of leaf protein is stored in sion during senescence and approximately the same number
the chloroplast but, perplexingly, the majority of the pro- are downregulated. The functions of some of these genes can
teolytic activity associated with senescing leaves appears to be be inferred from their annotation. Figure 3 illustrates the
in the vacuole. Recently, the importance of autophagy in the gene ontology (GO) annotations (http://www.geneonto-
senescence process has become evident with the identification logy.org) for the groups of up and downregulated genes and
of several senescence-enhanced genes in Arabidopsis encod- shows that many more regulatory factors are upregulated
ing orthologues of yeast autophagic genes. Autophagy is a including kinases, transcription factors and protein binding
common eukaryotic process by which cytoplasm and organ- factors. This illustrates the considerable regulatory activity
elles are taken up by the vacuole for degradation. Arabidopsis that is an essential feature of the senescence process. For
mutants in two autophagy genes show an accelerated senes- example, amongst the upregulated genes there are around
cence phenotype indicating that regulated autophagic activ- 80 putative transcription factors. In addition, there are
ity is essential for controlled senescence. In addition, the many genes encoding degradative functions, proteases, nuc-
occurrence of different types of vacuole has been observed in leases, cell wall and lipid degradation enzymes, etc. Unsur-
senescing leaf; a population of small lytic vacuoles develop in prisingly, a major group of the downregulated genes encode
the peripheral cytoplasm, close to the chloroplasts. These proteins involved in chloroplast development, chlorophyll
small vacuoles have been shown to contain the senescence- biosynthesis and photosynthetic processes. A further
specific protease SAG12, and are likely to have an important experiment recently undertaken in the authors lab, using
role in protein degradation during senescence. RNA isolated from 11 time points during leaf development
The amino acids released by protein degradation are has shown that there are multiple patterns of gene expres-
mobilized from the leaf and transported in the phloem. sion within this collection with groups of genes showing
There is some conversion of amino acids into glutamine different times of expression. Figure 2 illustrates two groups
and asparagine, the levels of these are disproportionately of genes showing opposite expression patterns; the protease
high in the phloem leaving a senescing leaf and both gluta- encoding gene SAG12 is in the group of upregulated genes
mine and asparagine synthetase transcript levels and while many genes involved in photosynthesis are in the
activity increase during leaf senescence. Conversion into group of downregulated genes. It is likely that genes in-
glutamine or asparagine is efficient since this allows the volved in the same degradative pathway will be co-regu-
transport of extra N moieties per molecule. lated and the mechanisms by which expression of such gene
clusters is controlled are currently being investigated with the
Other macromolecules long-term aim of identifying the key regulators of senescence.
Senescence can be induced artificially by dark treatment
Lipids are remobilized during senescence via a process of or by removal of the leaf from the plant. These treatments
b-oxidation and gluconeogenesis. Some carbon may be cause yellowing and protein degradation but are not nec-
mobilized from a senescing leaf but it is likely that most of essarily true representation of the natural process of
the lipid is used to provide energy for the other senescence developmental senescence. Recent microarray experiments
processes to occur since the level of energy supplied by comparing developmental senescence with artificially
photosynthesis is obviously reduced as senescence pro- induced senescence have indicated many common features
gresses. Nucleic acids, especially RNA, provide a valuable but also some significant differences. For example, the
source of mobilizable phosphorous and RNA levels drop signalling pathway involving the hormone salicylic acid is
rapidly as senescence enters the final stages. The nuclear important in developmental senescence but is not activated
DNA levels remain stable to the final stages of senescence in artificially induced senescence.
to enable de novo transcription capability to be maintained.

Regulation of Leaf Senescence

Gene Expression during Senescence
Hormonal regulation
Some time ago, it was shown that senescence is an active
process that requires new genes to be transcribed and Many plant hormones have been implicated as having a role
proteins to be synthesized. Since then, using a variety of in controlling leaf senescence and different plant species may
molecular techniques culminating in microarray analysis, have different hormone dependencies. In general, cytokinin

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 Senescence in Plants

250

200

150

100

50

0 Trans- Other
Transe- Other Trans- Hydro- Nucleo- DNA or Nucleic Recep-
Kinase Other cription mole- Protein
ferase enzyme porter lase tide RNA acid tor
activity binding factor cular binding
activity activity activity activity binding binding binding binding
activity function
sen 176 171 147 146 113 84 80 67 55 45 38 12 6
MG 141 217 81 144 60 71 25 57 44 56 18 12 15

Figure 3 Potential functions of genes up or downregulated during senescence. Gene Ontology (GO) annotations were applied to the groups of genes
that were either up or downregulated during senescence. Senescence enhanced genes (sen) are in pale green, while genes downregulated in senescence
(MG) are shown in dark green. Functional classifications are shown under each pair of cones and the numbers of genes in each group are shown in the table
below.

is seen as being a negative regulator of senescence since treat- Regulatory genes

ment of leaves with cytokinin delays the onset of senescence.
An elegant experiment using transgenic tobacco expressing Several potential regulatory genes have been identified by
a cytokinin biosynthesis gene from a senescence-enhanced map-based cloning from Arabidopsis mutants showing
promoter showed the importance of cytokinin very clearly. delayed leaf senescence. These include mutants in genes
At the onset of senescence, the promoter was induced and controlling the signalling pathways described above and
the synthesis of cytokinin occurred. This resulted in inhibi- also a small number of genes with unknown functions. For
tion of senescence and the leaves remained green. example, the ore9 gene encodes a protein with a potential
Hormones with a potential role in the induction of role in protein degradation, ore4 encodes a plastid rib-
senescence include ethylene, salicylic acid and jasmonic osomal subunit, etc. Recently, a gene encoding a NAC
acid. Levels of all these hormones increase during senes- domain transcription factor has been shown to be impor-
cence and different groups of senescence-enhanced genes tant for normal senescence. However, in all these mutants
depend on each pathway for expression. However, the role senescence is delayed but not stopped; leaves do eventually
and importance of each in the senescence process is not enter what appears to be a normal senescence process.
always clear. Mutants in ethylene signalling show delayed There is no information yet on what the role of these genes
leaf senescence but unlike fruit that depend on ethylene for is in the senescence process and no mutants have been
ripening, leaves lacking the ethylene signal do senesce identified that inhibit senescence totally. The reason for
eventually. Ethylene signal perception is age dependent in this is probably due to the plasticity of plant processes,
Arabidopsis; young leaves treated with ethylene are not other pathways can be switched on if there is a block in the
induced to senesce, but once a certain age threshold has usual sequence of events. Therefore, the key controlling
been reached, ethylene treatment results in the rapid genes of senescence remain to be identified.
induction of early senescence. Treatment of Arabidopsis
leaves with jasmonic acid (JA) results in the induction of
senescence but mutants in JA signalling do not show an Value of Senescence Research
obvious delayed senescence phenotype. All these three
hormones have strong links to stress signalling and the Senescence is the final stage of plant development and has a
importance of them in controlling gene expression during key role to play in generating sufficient reserves for the
senescence is indicative of the considerable cross talk that plant to survive, either in following seasons or in the next
exists between senescence and stress response pathways. generations. Cereal plants that have been bred to remain

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 Senescence in Plants

green longer have higher yields and increased tolerance to and signalling pathways between developmental and dark/starvation
stress. Premature senescence induced by stress results in induced senescence in Arabidopsis. Plant Journal 42: 7–585.
reduced yield and quality in crops. Post-harvest senescence Feller U and Fischer A (1994) Nitrogen metabolism in senescing leaves.
Critical Reviews of Plant Science 13: 41–273.
also has a severe implication on nutrient quality of both
Gan S and Amasino RM (1996) Cytokinins in plant senescence: from
forage crops and green vegetables. Thus, if we could iden- spray and pray to clone and play. Bioessays 18: 557–565.
tify and manipulate the key regulatory genes that control Gan S (2007) Senescence processes in plants. Annual Plant Reviews, vol.
senescence many important agronomic improvements 26. Ames, IA: Blackwell Publishing.
could be made that would be beneficial to growers and Hortensteiner S (2006) Chlorophyll degradation during senescence.
consumers. Annual Review of Plant Biology 57: 55–77.
Hörtensteiner S and Feller U (2002) Nitrogen metabolism and remobi-
lization during senescence. Journal of Experimental Botany 53:
Further Reading 927–937.
Lim PO, Woo HR and Nam HG (2003) Molecular genetics of leaf
Buchanan-Wollaston V, Earl S, Harrison E et al. (2003) The molecular senescence in Arabidopsis. Trends in Plant Science 8: 272–278.
analysis of plant senescence – a genomics approach. Plant Biotech- Thomas H and Stoddart J (1980) Leaf senescence. Annual Review of
nology Journal 1: 3–22. Plant Physiology 31: 83–111.
Buchanan-Wollaston V, Page T, Harrison E et al. (2005) Comparative Thomas H, Ougham H, Wagstaff C and Stead A (2003) Defining senes-
transcriptome analysis reveals significant differences in gene expression cence and cell death. Journal of Experimental Botany 54: 1127–1132.