Blackwell Science, LtdOxford, UKPCEPlant, Cell and Environment0016-8025Blackwell Science Ltd 2004? 2004
27911591168
Original Article
Branch autonomy under differential shading conditions
A. Lacointe
et al.
Plant, Cell and Environment (2004) 27, 1159–1168
Testing the branch autonomy theory: a 13C/14C
double-labelling experiment on differentially
shaded branches
A. LACOINTE1, E. DELEENS2,†, T. AMEGLIO1, B. SAINT-JOANIS1, C. LELARGE2, M. VANDAME1, G. C. SONG3 &
F. A. DAUDET1
1
U.M.R. PIAF (INRA – University Blaise Pascal), Site INRA de Crouelle, 234 av. du Brezet, 63039 Clermont-Ferrand Cedex
2, France, 2I. B. P., bâtiment 630, Université Paris-sud, 91405 Orsay, France and 3National Horticultural Research Institute,
Rural Development Administration, Suwon 440–310, South Korea
ABSTRACT
The impact of a heterogeneous within-crown light environment on carbon allocation was investigated on young walnut trees trained on two branches: one left in full sunlight,
the other shaded until leaf fall resulting in 67% reduction
in photosynthetically active radiation. In September, the
two branches were separately labelled with 14CO2 and
13
CO2, respectively, so that the photosynthates from each
branch could be traced independently at the same time.
Although some carbon movements could be detected
within 5 d in both directions (including from the shaded
branch to the sun branch), between-branch carbon movements were very limited: approximately 1% of the diurnal
net assimilation of a branch. At this time of the year branch
autonomy was nearly total, leading to increased relative
respiratory losses and a moderate growth deficit in the
shaded branch. The ratio of growth to reserve storage rate
was only slightly affected, indicating that reserves acted not
as a mere buffer for excess C but as an active sink for
assimilates. In winter, branch autonomy was more questionable, as significant amounts of carbon were imported
into both branches, possibly representing up to 10% of total
branch reserves. Further within-plant carbon transfers
occurred in spring, which totally abolished plant autonomy,
as new shoots sprouted on each branch received significantly more C mobilized from tree-wide reserves than from
local, mother-branch located reserves. This allowed great
flexibility of tree response to environment changes at the
yearly time scale. As phloem is considered not functional
in winter, it is suggested that xylem is involved as the pathway for carbohydrate movements at this time of the year.
This is in agreement with other results regarding sugar
exchanges between the xylem vessels and the neighbouring
reserve parenchyma tissues.
Correspondence: A. Lacointe. Fax: +33 (0)4 73 62 44 54; e-mail:
[email protected]
†
Eliane Deléens passed away on 13 March 2003.
© 2004 Blackwell Publishing Ltd
Key-words: allocation; balance; carbohydrate; carbon;
mobilization; reserve; tree.
INTRODUCTION
Among the environmental factors that affect tree growth
and development, light plays a central role as the major
factor involved in photosynthesis. Whole tree shading or
canopy opening experiments have shown that shading
results in decreasing, not only total growth, but also the
root/shoot ratio (Van Hees 1997; Tognetti et al. 1998). Shading generally affects more readily radial growth than primary growth (Kappel & Flore 1983; Collet, Lanter &
Pardos 2001), and may affect flowering and/or fruiting
(Ryugo, Marangoni & Ramos 1980; Proctor & Crowe 1983;
Marini & Sowers 1990).
Inside tree crowns, local light conditions can be very
different among branches, so that the general effects of the
shading level as mentioned above can be expected to result
in differential growth and development patterns within the
crown. This assumes, however, that the direct effect of light
on photosynthesis – and hence on the local carbon balance
between production and demand for growth and respiration – is not compensated for by photo-assimilate
exchanges between branches of different light status. The
latter assumption, called the ‘branch autonomy’ theory
(Sprugel, Hinckley & Schaap 1991; Brisson 2001), has been
invoked as a mechanism for differential branch growth and/
or survival according to local light conditions, which in turn
can explain specific tree shapes in relation to stand density
or age. The ‘branch autonomy’ concept has been included
in several tree growth models (Ford, Avery & Ford 1990;
Takenaka 1994; Kellomäki & Strandman 1995).
To this day, experimental evidence in support of the
branch autonomy theory has been mostly indirect, through
its expected consequences. Defoliating or shading individual branches of pine (Honkanen & Haukioja 1994) or birch
(Ruohomaki et al. 1997; Henriksson 2001) significantly
affected their mid- or long-term growth or survival rate,
often more severely for a branch-wide than for a tree-wide
treatment. Looking at carbohydrate reserve dynamics in
1159
1160 A. Lacointe et al.
individual prune branches, Ryugo et al. (1977) reported at
least partial autonomy of fruiting versus non-fruiting
branches through local reserve mobilization; similarly,
Langstrom et al. (1990) reported a significant decrease in
starch reserves of partly defoliated individual pine
branches whereas reserves in neighbouring branches were
not affected.
However, direct experimental evidence through tracing
photo-assimilate movements between branches of different
light status is surprisingly limited: we are aware of only two
investigations in this area. Using 14C labelling, Cregg, Teskey & Dougherty (1993) concluded that ‘loblolly pine
shoots were usually autonomous with respect to carbohydrate supply, but that carbohydrate movement into the terminal shoot from subtending foliage could occur under
conditions of very high stress’ [i.e. high growth demand].
However, only mature needles, excluding growing organs
and mature stem tissues, had been evaluated as possible
sinks for between-branch transfers, so that conclusion
should be considered partial. In the other experiment, on
persimmon, Yamamoto et al. (1999) showed that ‘a very
small quantity of 13C photosynthates was exported into
neighbouring lateral branches for 44–72 h after a basal or
terminal lateral branch was exposed to 13CO2’; unfortunately, due to individual variability the results were not
conclusive about a possible effect of shading.
The first objective of the present investigation was to
increase experimental knowledge about assimilate movements between differentially shaded branches. In order to
evaluate the specific effects of shading, reciprocal transfer
rates between a sunlit and a shaded branch were measured
simultaneously in the same trees using double 13C/14C labelling, on the simplest possible experimental system: tree
crowns consisting of only two branches.
Furthermore, beside current photo-assimilates soluble
sugars can be found in the xylem sap of a number of species
such as maple or walnut in winter (Améglio & Cruiziat
1992; Améglio et al. 2001), and at budbreak growth resumption can mobilize up to 50% of the tree’s total carbon
reserves (Lacointe et al. 1993, 1995a; Barbaroux, Bréda &
Dufrêne 2003). The scope and significance of these potentially large scale movements with respect to branch autonomy was addressed by periodic sampling.
MATERIALS AND METHODS
Plant material
Sixteen 2-year-old Juglans regia L. scions, cv. ‘Franquette’,
were planted in 33 L containers. The soil was a peat : clay
mixture (33% : 67%; vol : vol) complemented with 10 g
NH4NO3, drip irrigated to field capacity. In April, the stem
was partially disbudded, allowing only two branches to
grow out. In early July, the lower branch was shaded with
a shading net allowing approximately 33% of incident
PAR, which resulted in a 30% decrease of its diurnal net
assimilation. A few trees were kept in full sunlight as
controls.
Isotopic labelling and sampling
In mid-September, each branch was separately, quantitatively labelled with *CO2 in 40 L chambers (for details see
Kajji et al. 1993). The sunlit branch (open circle) was fed with
18 MBq 14CO2, the shaded one (closed circle) with 4 mmoles
13
CO2. The shading system was removed at leaf shedding
(November). Three trees were harvested at each of the
following dates (four dates and 12 trees): 5 d after labelling
(end of primary translocation, cf. Lacointe et al. 1995b); leaf
shedding; just prior to budbreak (next spring); and in June,
when new shoots displayed four fully grown leaves.
Biochemical analyses
After harvest, the plants were rapidly divided into coarse
roots, fine roots, main stem, branches and new shoots when
present, and immediately frozen in liquid nitrogen. After
freeze-drying, each organ was ground to pass a 125-mm
mesh. Soluble sugars were extracted in 80% ethanol at
80 ∞C, then purified successively on active charcoal + polyvinyl-polypyrrolidone (PVPP), cation exchanger and anion
exchanger resins. Glucose, fructose and sucrose were
assayed by the enzymatic method of Boehringer (1988).
After enzymatic extraction from the pellet, starch was
quantified as glucose equivalent (Boehringer 1988). The
sum of soluble sugars plus starch is hereafter referred to as
‘mobilizable carbohydrate’ fraction, whereas the nonextractable fraction will be considered ‘structural carbon’.
Isotopic analyses
The 14C of the whole dry matter was determined at ‘infinite
thickness’ in a low-background argon–methane flow
counter (Numelec NU 20; Canberra Eurisys, SaintQuentin-en-Yvelines, France). The 14C contents of the
purified sugar extract and of the extracted starch were measured on all organs by liquid scintillation. For 13C, the total
dry matter label was determined on all organs using an
elemental analyser (Carlo Erba NA 1500; Thermo Electron
Corporation, Milan, Italy) connected to an isotope mass
spectrometer (Optima; Fisons, Villeurbanne, France); the
13
C atom percentage excess for each organ was computed
versus an unlabelled control tree. The same device was also
used to determine the 13C content of soluble carbohydrates
and of starch in each branch, after the same extraction
procedure as for 14C, with a further purification step on
PVPP + ion exchanger resins for starch extraction.
For both isotopes, the excess label recovered in each
organ ¥ biochemical fraction was expressed as ‘unit label’:
1 unit label = 1 per-cent (%) of the total amount of label
initially fed to the tree. The resulting value (L) was then
corrected for individual tree and organ size variability (and
associated dilution effects) to yield a normalized value:
Normalized label (NL)
whole tree SM ¥ average organ SM
=L¥
organ SM ¥ average whole tree SM
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
Branch autonomy under differential shading conditions 1161
where ‘SM’ stands for perennial structural dry matter mass
(g), and ‘average’ applies to all trees harvested at a given
date.
In organs that lost some carbohydrates between the two
sampling dates, the specific label (unit label per g carbohydrate) of the mobilized carbohydrates was estimated as the
ratio of carbohydrate NL variation to the variation of ‘normalized carbohydrate amount’ (NC)
average organ SM
NC = amount of compound in organ ¥
organ SM
Results are expressed as average ± standard error values.
Unless otherwise stated, significance of differences is
assessed by Student’s t-test (two-tailed, heteroscedastic
version).
Table 2. Spatial partitioning of the carbon (‘normalized label’,
see Material and methods) exported from each source branch in
September, expressed as percentage of the total exported C,
namely the % of the total recovered outside of the source branch
Source branch
Opposite branch
Main stem
Roots
Sunlit ()
Shaded ()
0.17 ± 0.04*
21 ± 1
80 ± 1
0.27 ± 0.08*
22 ± 4
78 ± 4
*Value significantly > 0 (P < 0.05, Student’s t-test, one-tailed
distribution).
Carbon allocation in September
RESULTS
Effect of light on branch growth and
carbohydrate content
The sunlit branch exhibited a slightly (but significantly:
P < 0.05, non-parametric Wilcoxon paired test) higher total
dry matter mass (DM) than the shaded one (Table 1). However, for practical reasons the sunlit branch was also the
upper one whereas the shaded branch was in a lower position within the tree architecture, so that it was not clear
which factor this effect was due to. To address this question,
full-sun-grown trees were also investigated as controls. No
such difference was found between the two branches of
these control trees (g DM: 12 ± 2 for the upper branch versus 11 ± 2 for the lower one; P = 0.78, same test as above),
so that the higher DM of sunlit versus shaded branches
could actually be ascribed to the differential light environment between both branches rather than to their respective
position within the tree architecture.
Regarding carbohydrate concentration, no difference
was detected between both branches (Table 1), although
the difference in total DM resulted in a similar difference
in total (absolute) carbohydrate content between
branches.
A total of 52% of the total C assimilated by the shaded
branch was lost from the tree within 5 d after labelling
(Fig. 1a), versus only 35% for the sunlit branch (significance level of difference: P = 0.01). For both isotopes, less
than 40% of the total label that was recovered within the
source branch was incorporated in the ‘residual’ noncarbohydrate fraction, which is an indicator for anabolic
activity-like growth, although the proportion was slightly
higher in the shaded than in the sunlit branch (Fig. 1b).
However, the spatial partitioning patterns were remarkably
similar: in both cases, approximately 60% of the total C
recovered in the whole plant was exported out of the source
branch, the other 40% remaining within it. The patterns of
partitioning among sink organs (Table 2) were also identical, as approximately 80% of the exported C was allocated
to roots versus 20% allocated to the main stem, which was
not surprising as they both reflected the same process
(assimilate partitioning among the main sink organs),
revealed by two independent tracers. However, the most
interesting result was that from each source branch, regardless of light environment, a low but statistically significant
proportion (approximately 0.2%) of the exported C was
recovered in the opposite branch.
Autumn and winter dynamics
Table 1. Dry matter mass (DM) and carbohydrate content of the
different parts of the trees at leaf shedding, in November
Organ
Total DM
(g)
Carbohydrates
(mg g-1 DM)
Sunlit branch ()
Shaded branch ()
Main stem
Coarse roots
Fine roots
13 ± 2*
9 ± 1*
60 ± 10
200 ± 30
40 ± 15
90 ± 5
88 ± 3
108 ± 1
412 ± 10
147 ± 7
Carbohydrates (soluble sugars + starch) are expressed as mg
glucose equivalent per g DM. * : DM ratio is greater than 1
(non-parametric Wilcoxon significance level: a < 0.05)
The sunlit branch-derived carbon incorporated in the structural fraction of dry matter exhibited no change in the roots
or the main stem in autumn or winter (Table 3). However,
there was an increase in both branches, particularly
between November and April (significance levels: P = 0.08
for the source branch; P = 0.05 for the opposite branch).
At the same time, the labelled carbohydrates decreased
in most organs, particularly between September and
November. However, the opposite (= shaded) branch did
not fit in this pattern, as this was the one organ in which
labelled carbohydrates did not decrease, but increased significantly, in the same way as labelled structural carbon,
both in autumn (September to November: P = 0.01) and in
winter (November to April: P = 0.01). When added, both
fractions (i.e. the total dry matter label) in this branch
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
1162 A. Lacointe et al.
Figure 1. Carbon allocation pattern in
September, 5 d after assimilation. (a)
Within-tree spatial distribution: for each
source branch (, sunlit branch; , shaded
branch), the figure shows the percentage of
total photosynthates (‘normalized label’,
see Material and methods) that are recovered either within the source branch (in
grey) or in the rest of the tree (export, in
black). Assimilates not recovered in either
compartment are ascribed to respiratory
losses. (b) Chemical partitioning of total
carbon label recovered within each source
branch, 5 d after assimilation in September: , sunlit branch (14C); , shaded
branch (13C). Vertical bars represent standard errors.
exhibited a very significant increase of approximately 100%
(representing approximately 0.075 unit label in absolute
terms) between November and April (P = 0.01).
In the different organs that lost some labelled carbohydrates between November and April, and were thus potential carbon sources for the concomitant label increase in the
shaded branch, the specific label of mobilized carbohy-
drates was approximately 0.4 label units per gram glucose
equivalent as an order of magnitude (Table 4). This provides a rough estimate of the (minimum) amount of carbohydrate that had been mobilized and translocated into the
shaded branch from other parts of the plant: 0.035/
0.4 = 0.1 g carbohydrate as an order of magnitude.
Furthermore, both branches appeared symmetrical in
Table 3. Autumn and winter variations of sunlit branch-derived carbon in the extractable carbohydrate (= starch + soluble sugars) and the
non-extractable (= ‘structural’) fractions of each part of the tree (excluding leaves in September)
NL in non-extractable structures
September
Source branch
Opposite branch
Main stem
Coarse roots
Fine roots
Total
1.0 ± 0.1
0.015 ± 0.004
2.6 ± 0.3
5.4 ± 1.2
6.8 ± 0.7
16 ± 1
NL in mobilizable carbohydrates
November
*
1.6 ± 0.2
0.05 ± 0.01
2.6 ± 0.4
5.5 ± 1.1
8±2
18 ± 3
*
April
September
November
April
2.5 ± 0.3
0.09 ± 0.01
2.5 ± 0.1
4.5 ± 1.2
8±1
2.6 ± 0.1
0.016 ± 0.001
5.6 ± 0.5
14 ± 2
4.8 ± 0.7
*
*
*
†
†
1.2 ± 0.2
0.035 ± 0.003
2.0 ± 0.5
8.6 ± 2.1
2.8 ± 0.8
†
†
0.9 ± 0.2
0.070 ± 0.006
1.5 ± 0.1
5.8 ± 1.4
2.1 ± 0.3
18 ± 2
27 ± 3
*
14 ± 3
*
10 ± 2
*
Data are expressed as normalized label (NL) values (cf. Material and methods). * The difference between adjacent columns is significant
(P = 0.05, Student’s t-test). † The difference between September and April was significant (P = 0.05), although those between September
and November and between November and April were not.
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
Branch autonomy under differential shading conditions 1163
Table 4. Estimated specific label of 14C-carbohydrates mobilized
in the whole tree between November and April (Normalized values, cf. Material and methods)
Label of mobilized carbohydrates (unit label)
Amount of mobilized carbohydrates (g glucose
equivalent)
Specific label of mobilized carbohydrates
(unit label g-1)
4±3
16 ± 3
0.4 ± 0.3
this respect (Fig. 2), although due to a higher error in 13C
than 14C excess evaluation, the increase in 13C was not statistically significant (P = 0.22).
Spring mobilization
Between November (leaf fall) and June (new shoots
became self-sufficient), 75% of the total tree mobilizable
sunlit branch-derived carbohydrates were used up (Fig. 3).
Of that carbon, approximately 40% was recovered in the
new shoot dry matter. However, most of the total consumption of labelled carbohydrates occurred in spring, between
budbreak and new shoot self-sufficiency. This was massive
mobilization, as 67% of the labelled reserves still present
in April were lost in June, with approximately 62% of that
spring-mobilized carbon recovered in new shoots.
Interestingly, the new shoots sprouted on the unlabelled
branch got a much higher amount of label than expected
from the pre-mobilization label content of their mother
branch, and the same was true for the shaded branchderived carbon (Table 5). As a result, in the sunlit branchborne new shoots, the ratio of shaded branch-derived C
(13C) to sunlit branch-derived C (14C) was increased by a
factor of 9 as compared to their mother branch. A symmetric situation was observed in the shaded branch-borne new
shoots versus their mother branch, although the factor was
only 3.
This dilution of ‘local reserve originating C’ by ‘wholetree reserve originating C’ in new shoot dry matter could
be viewed in a more quantitative way through a simple
modelling approach, as the experimental design by double
labelling provided two independent mobilization patterns,
which allowed model fitting and model evaluation on two
independent data sets.
A simplified model of spring mobilization and
allocation of whole tree reserves to branches
This model was developed with the purpose of gaining a
better understanding of the limiting factors and mechanisms involved in reserve mobilization associated with new
shoot growth.
The total label of new shoots sprouted from each branch
in June (NSL) is computed as the sum of a ‘global’ and a
‘local’ component. The ‘global’ component is assumed proportional to total label of carbohydrate reserves in main
stem and coarse roots in April (global reserve label: GRL),
Figure 2. Winter variations of the label
derived from each branch that is recovered
in the opposite branch. Total label is partitioned into extractable carbohydrates (=
starch + soluble sugars) and non-extractable (= ‘structural’) carbon. Data are
expressed as normalized label (NL) values
(cf. Material and methods). (a) Sunlit
branch-derived carbon recovered in the
shaded branch; (b) shaded branch-derived
carbon recovered in the sunlit branch. Vertical bars represent standard errors.
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
1164 A. Lacointe et al.
14.5
± 3.3
Figure 3. Winter and spring mobilization
of sunlit branch-derived carbohydrate
reserves: seasonal course of total reserves
of whole tree (excluding new shoots in
June), and reserve-derived carbon recovered in total dry matter of new shoots in
June. Data are expressed as normalized
label units (NL, cf. Material and methods).
10.4
± 1.6
3.5
± 0.6
possibly multiplied by new shoot mass (NSM) to account
for a possible effect of new shoot growth rate. The ‘local’
component is assumed proportional to the label of carbohydrate reserves in bearing shoot in April (local reserve
label: LRL), also possibly multiplied by new shoot mass
(NSM).
As a first step, fitting was performed on the 14C data set,
with the purpose of finding the best version of the model,
constrained by the condition that the local component must
not exceed the total amount of label actually mobilized
from the bearing shoot. Least-mean-squared errors of fitting (MSE) – or highest coefficient of determination (R2),
which is formally equivalent but easier to read— was used
as the fitting criterion.
As expected, the above-mentioned predominance of
‘global’- over the ‘local’-originating C appeared as a higher
contribution to goodness-of-fit of the global component,
provided it included the NSM proportionality (Table 6a):
in single-component versions of the model, it could account
for over 80% of total as compared to approximately 60%
for the local one alone. Furthermore, the specific contributions of both components appeared relatively independent
as the full model accounted for over 90% of total
variability.
Although NSM (new shoot mass, i.e. cumulative growth)
clearly contributed to the significance of the global component, this first step yielded no information about the significance of the global reserve store itself, as there was only
one GRL value available in the 14C data set. This could be
discriminated in the second step, the evaluation procedure,
which was performed on the 13C data set, with the least
mean squared error of prediction (MSEP, computed using
the parameters resulting from the fitting step) as the evaluation criterion.
The evaluation (Table 6b) confirmed the significance of
the local component beside the global, as it reduced MSEP
by a factor of more than 2. The most interesting point,
however, was that the best model (least MSEP) led to a
relative error of prediction (the square root of MSEP
divided by mean experimental value) that was less than
30%, i.e. only 1.5 times as high as the average relative error
of fitting (the square root of MSE divided by mean experimental value). The corresponding model version involved
NSM in the global but not in the local component, suggesting that the mobilization of local reserves could be less
dependent on new shoot growth than that of global
reserves.
DISCUSSION
Effect of shading on branch growth and fate of
current photo-assimilates
Consistent with previous works on different species (Honkanen & Haukioja 1994; Ruohomaki et al. 1997; Henriksson 2001), differential shading had a slight but statistically
significant effect on growth, as roughly measured by total
dry matter. The use of isotopic tracers revealed another,
major, effect: the carbon losses from the shaded branchassimilated carbon were significantly higher than those
from the sunlit branch-assimilated C. At this time scale, i.e.
within 5 d after assimilation, carbon losses can be ascribed
to respiration (Lacointe et al. 1995b), so this denotes higher
relative respiratory losses in the former than in the latter
case. As the metabolic requirements of the shaded branch
can be assumed slightly lower than, or at most similar to,
those of the sunlit branch, this is an indication for a lower
absolute amount of available carbohydrate substrate, as
expected if the lower photosynthetic production was not
compensated for by import from other tree parts. However,
Table 5. Total label of carbohydrate reserves in both bearing branches just prior to budbreak (April) and of total dry matter in new shoots
derived from each bearing branch a few weeks after budbreak (June)
13
Total carbohydrates in
bearing branch (April)
Total DM in new shoots sprouted
from bearing branch (June)
C
0.06 ± 0.02
0.6 ± 0.1
1.9 ± 0.2
2.1 ± 0.2
14
C
0.9 ± 0.2
0.07 ± 0.01
3.4 ± 0.4
0.8 ± 0.2
ratio 13C : 14C
0.07 ± 0.04
9±2
0.6 ± 0.1
3±1
Symbols:, ,bearing branch grown in full sun light and fed with 14C in the previous year; , bearing branch shaded and fed with 13C in the
previous year. Data are expressed as normalized label units (cf. Material and methods).
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
Branch autonomy under differential shading conditions 1165
Table 6. Parameterization and evaluation of the model: NSL = Global + Local, where: NSL = total label of new shoots sprouted from each
branch in June (norm. label units); Global = one of the following: {0, b, b ¥ GRL, b ¥ NSM, b ¥ NSM ¥ GRL}; Local = one of the following:
{0, a ¥ LRL, a ¥ NSM ¥ LRL}; with: a, b = constant parameters; NSM, total dry matter mass of new shoots (g); LRL, ‘local reserve
label’ = total label of carbohydrate reserves in bearing branch in April (normalized label units); GRL, ‘global reserve label’ = total label of
carbohydrate reserves in main stem + coarse roots in April (normalized label units). The model is constrained by the condition: ‘Local < total
label actually mobilized from bearing shoot’.
(a) Parameters of the model as fitted on data relative to the mobilization of C originating from the sunlit branch, i.e. the 14C data. R2,
coefficient of determination.
Local component
a ¥ LRL
0 (none)
a ¥ NSM ¥ LRL
Parameter value
Parameter value
Parameter value
Global
component
R2
a
b
R2
a
b
R2
a
b
0 (none)
b (= constant)
b ¥ GRL
b ¥ NSM
b ¥ NSM ¥ GRL
–
0
0
0.84
0.84
–
–
–
–
–
–
2.1
0.28
0.072
0.010
- 1.33
0.40
0.40
0.92
0.92
0.69
0.69
0.69
0.69
0.69
–
1.7
0.24
0.059
0.0081
- 0.75
0.62
0.62
0.97
0.97
0.026
0.026
0.026
0.026
0.026
–
1.5
0.21
0.052
0.0071
(b) Evaluation on 13C data, of the best versions of the model as fitted on 14C data, i.e. with parameter values from Table 6a. The evaluation
criterion is MSEP, mean squared error of prediction (squared normalized label units, cf. Materials and methods). For comparison, the table
also provides mean squared errors of fitting (MSE, same units as MSEP), which are equivalent to R2 values in Table 6a.
Local component
a ¥ LRL
0 (none)
a ¥ NSM ¥ LRL
Global
component
MSE (14C)
MSEP (13C)
MSE (14C)
MSEP (13C)
MSE (14C)
MSEP (13C)
b ¥ NSM
b ¥ NSM ¥ GRL
0.28
0.28
0.92
0.72
0.13
0.13
0.36
0.28
0.06
0.06
0.36
0.38
the most direct evidence in support of branch autonomy
was of course the very low level of between-branch photoassimilate movements: from each source branch, regardless
of light environment, only approximately 0.2% of the total
exported C was recovered in the opposite branch. It can be
calculated that this represented approximately 1% of the
branch own diurnal net assimilation. Thus, it can be concluded that branch autonomy was nearly total regarding
primary assimilate allocation.
In addition to the above-mentioned change in relative
allocation to respiration, shading had another detectable
effect on local carbon economy: the relative incorporation
into the ‘structural’ fraction was slightly higher, which can
be understood as a slight increase in the relative allocation
of C to growth versus reserve storage activity. This change,
however, was of limited extent, not significantly affecting
the final relative carbohydrate content of the branch DM
at the end of the season. In other words, the assimilate
shortage affected both the growth and reserve storage
rates, the latter being only slightly more affected. A similar
‘balanced covariation’ of both activities in response to different shading conditions was previously reported in young
oak (Ziegenhagen & Kausch 1995) and beech (Gansert &
Sprick 1998). Thus, although reserves certainly can act as
short- or middle-term buffers to cope with temporary
imbalance between C production and demand, such as during peak fruit growth (Ryugo et al. 1977), they should in
many cases not be considered as mere passive buffers in the
long term. Instead, they appear as a vital component that
the tree will (if possible) ‘manage’ to keep above a critical
level, possibly at the expense of growth in case of source
limitation. As a practical consequence, source–sink based
models of C partitioning in trees would likely benefit from
assigning reserve storage a competitive ‘sink strength’, as is
generally the case for growth (Lacointe 2000; Le Roux et al.
2001).
Autumn and winter dynamics
From 5 d after assimilation through leaf fall and winter, the
total label incorporated in the structural fraction exhibited
no change in most tree parts, namely the roots or the main
stem. This is consistent with previous results (Lacointe et al.
1995b) showing that leaf export, which was the limiting step
for the final pattern of spatial and biochemical partitioning
of current assimilates, was completed by 90% within 5 d. In
the present experiment, however, there was an increase in
the structural fraction of branches, both between
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
1166 A. Lacointe et al.
September and leaf fall, and even more after leaf fall. As it
could in neither case be ascribed to current assimilate
import and metabolism, this is an indication that some
later anabolic activity occurred in current-year shoots,
not only in the autumn but also in winter. Although this
experiment provided no qualitative information in this
respect, it might involve phenolic compounds, particularly
phenol glycosides, which have been found in particular
abundance in walnut tissues (Claudot, Drouet & JayAllemand 1992).
Regarding extractable carbohydrates, their total label
decreased in most organs between September and November This is an indication for turnover, suggesting a role as
mobilizable carbon source for the above-mentioned, postprimary export increase in source branch structural matter.
However, as shown by the labelled carbohydrate increase
in the opposite branch, part of that recirculating mobilized
C was also re-deposited as carbohydrate reserves at a distance from the place of remobilization. When both fractions
were added (extractable carbohydrates + structural C), the
increase in the opposite branch total dry matter label
exceeded 100%, so that branch autonomy before leaf fall
did not appear so strict as it did when considering only
primary assimilate partitioning.
Moreover, further and even more significant amounts of
mobile carbon were imported into both branches during
the winter, between November and April The amount of
carbohydrate that has been mobilized and translocated into
the shaded branch from other parts of the plant could be
roughly estimated as 0.1 g carbohydrate as an order of magnitude, which is very low (approximately 0.1%) compared
with the total tree reserves, but may be significant (approximately 10%) relative to the branches’ own reserves.
Although this figure was yielded by a very rough calculation, it should be considered rather underestimated as it
assumed that all of the imported carbon was recovered at
harvest, thus ignoring any respiratory losses.
An interesting question is the pathway for carbohydrate
movements in winter. As phloem is considered not functional in winter (Aloni 1991; Aloni & Peterson 1997), this
suggests the involvement of the xylem pathway. This is in
agreement with other results on walnut regarding sugar
exchanges between the xylem vessels and the neighbouring
reserve parenchyma tissues, with major consequences on
the water status and likely spring development (Améglio
et al. 2001; 2004).
Spring mobilization
A total of 75% of the total September-labelled reserves in
the whole tree were used up between November (leaf fall)
and June (new shoots became self-sufficient), with most of
the total consumption of labelled carbohydrates occurring
in spring, between budbreak and new shoot self-sufficiency.
This is consistent with the mobilization rates of Augustlabelled reserves (45%) and October-labelled (80%) as
previously found by Lacointe et al. (1993) in young walnut.
Of the total mobilized carbon, approximately 40% was
recovered in the new shoot dry matter, which again is consistent with the recovery rates of C derived from Augustlabelled reserves (60%) and October-labelled (15%) as
found by Lacointe et al. (1993).
Regarding the branch autonomy issue, however, the
most interesting point was that the new shoots sprouted on
each branch got a much higher amount of C initially
labelled in the opposite branch than expected from the premobilization label content of their own mother branch. This
resulted in significant dilution of ‘local reserve originating
C’ by ‘whole-tree reserve originating C’ in new shoot dry
matter. At this point, very little was left of branch
autonomy.
This dilution effect could be simulated by a simple model
of the respective contribution of ‘local’ (mother-branch)
versus ‘global’ (tree-wide) reserves. After fitting on the 14C
data set, the model fairly well simulated the 13C data set,
which can be considered not a mere replication but an
independent experiment in a different situation (regarding
the light environment at labelling). The results suggested
that the mobilization of ‘local’ (mother-branch) reserves
could be less dependent on new shoot growth than that of
‘global’ (tree-wide) reserves, which would be consistent
with a rather ‘source-limited’ mobilization for local
reserves whereas the tree-wide mobilization would be more
‘sink-limited’ or ‘sink-driven’. If so, the early stages of
mobilization just before or at budbreak can be expected
to involve preferentially ‘local’ reserves whereas the
‘global’ would be tapped on at later stages, in relation to
actual growth rate; this hypothesis could be tested by
frequent harvesting around and within a few weeks after
budbreak.
Taken all together, these results do not support the idea
of long-term branch autonomy, thus contradicting conclusions from previous defoliating or shading experiments,
particularly in birch, as reported in the Introduction section. However, the contradiction may be only apparent,
with at least three possible levels of explanation.
Firstly, some genetic factors, specific to each species,
might be involved. Although there is a balance between
growth and reserve storage in many species, including walnut, oak or beech as discussed above, this might not be the
case in pioneer species such as birch. If growth indeed has
priority over reserve storage as a sink in that species, local
shade would lead to local significant reserve deficit (which
did not occur in the present experiment in walnut). As a
consequence, early bud growth might be affected, in turn
hindering the subsequent growth-sink driven massive
import of tree-wide originating resources as hypothesized
above, etc., eventually leading to branch death as predicted
by the branch autonomy principle.
Secondly, it should be emphasized that this experiment
was carried out on young trees. However, as suggested by
a few investigations into the response to pruning with
respect to reserve dynamics (e.g. Clair-Maczulajtys & Bory
1988), within-tree C fluxes tend to become more ‘compartmentalized’ as trees grow older and larger. This would
mean a higher significance of local versus tree-wide
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168
Branch autonomy under differential shading conditions 1167
resources for local growth, resulting in better validity of
branch autonomy in old than in young trees.
Thirdly, although shading systems were removed in winter both in Henriksson’s (2001) and in the present experiment, light conditions were very different in spring. In the
former experiment, branches were re-shaded at leaf flush.
In contrast, in our experiment the shading system was not
re-installed, so that not only bud break but also subsequent
new shoot growth occurred in a sunlit environment, as
would occur after a clearing. This could explain the
observed difference, if the fate of resources from massive,
tree-wide reserve mobilization is, directly or not, dependent on the local environment. Assuming sink-driven
import, such dependence would arise from early sink
demand of new shoots, or early growth rate, which could
be stimulated by light through its own photosynthesis, or
maybe through more qualitative effects on development.
Through such a feedback loop, a clear light environment
would not only promote import into sunlit branches, but
also inhibit import of tree-wide originating resources into
shaded branches, in relation to competition among sinks.
This would further explain a seemingly paradoxical effect
observed by Henriksson (2001), which true branch autonomy would not allow: the impact of shading, as evaluated
by eventual death rate, was more pronounced on branches
that were individually shaded, hence competing with sunlit
ones, than on completely shaded trees.
CONCLUSIONS
In this experiment on young walnut, a heterogeneous
within-crown environment yielded contrasting results
regarding branch autonomy:
In summer, although some carbon movements could be
detected in both directions (including from the shaded
branch to the sunlit branch), between-branch carbon movements were very limited, being approximately 1% of the
diurnal net assimilation of a branch. Thus, at this time of
the year, branch autonomy was nearly complete, leading to
increased relative respiratory losses and a moderate growth
deficit in the shaded branch.
In winter, however, significantly more carbon was
imported into the branches, probably representing up to
10% of total branch reserves, which makes branch autonomy more questionable at the seasonal time scale. It was
even more so when including spring outgrowth, as new
shoots sprouted from each branch got more carbon mobilized from whole tree-wide reserves than from the local
reserves located in their own mother branch. However,
there were indications that this massive import of tree-wide
resources into the lateral branches in spring might be
dependent on the local light environment at that time.
In this view, the dynamics of tree carbon economy and
growth exhibits a great flexibility of response to environmental changes, as new shoots are granted a ‘new chance’
in spring if the environment is improved, as happens after
a clearing or a storm, even if the bearing shoot was previously under adverse conditions.
ACKNOWLEDGMENTS
This work was supported by the EU Project Fair CT961887 (W-BRAINS), with Jean-Sylvain Frossard as the local
co-ordinator. Additional technical assistance was provided
by Christian Bodet, Maurice Crocombette and Stéphane
Ploquin. We thank Pascale Maillard for helpful discussions.
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Received 16 March 2004; received in revised form 5 May 2004;
accepted for publication 21 May 2004
© 2004 Blackwell Publishing Ltd, Plant, Cell and Environment, 27, 1159–1168