Discussion
Light-responsive photosynthate translocation to
fruits
Photosynthate translocation is directly or indirectly influenced by
environmental factors such as light intensity, air temperature, drought
stress and CO2 concentration. In protected horticulture,
it is important to understand the response of photosynthate
translocation to different environments in order to establish an
environmental control system supporting high yield and high-quality
fruit production. Many studies have therefore assessed the relationship
between different environmental factors and dynamics of photosynthate
translocation. Regarding air temperature, Yoshioka (1986) reported an
increased translocation speed of photosynthates with an increasing
temperature in petioles of tomato plants. Pickard & Minchin (1990)
showed the inhibition of phloem translocation by abrupt 10°C temperature
drops in the stems of Phaseolus vulgaris . Makino & Mae (1999)
reviewed the effects of elevated CO2 levels on
photosynthesis and plant growth. It was found that short-term
CO2 enrichment stimulated the rate of leaf
photosynthesis and enhanced plant biomass. In contrast, prolonged
exposure to CO2 enrichment generally reduced the initial
stimulation of photosynthesis and often suppressed leaf photosynthesis
by means of secondary responses (e.g. accumulation of carbohydrates,
decrease in leaf nitrogen and Rubisco concentration). Sung & Krieg
(1979) described a reduction in sensitivity of photosynthetic
assimilation and translocation to drought stress in leaves of cotton and
sorghum. Furthermore, several studies describe the relationship between
photosynthate translocation and light conditions because of their direct
link through photosynthesis (Troughton et al. 1977; Lemoineet al. 2013; Lanoue et al. 2018) since light provides
energy for the synthesis of sugars. Troughton et al. (1977) and
Lanoue et al. (2018) evaluated the effects of light intensity and
light quality on photosynthate translocation in leaves of maize and
tomato, respectively, by using 11C- and14C-labelled CO2, respectively. Both
of these studies found the straightforward result that photosynthate
translocation increases as light intensity increases. Whereas all of the
above-mentioned studies reported the influence of several factors on the
photosynthate translocation within stem and leaf parts, we investigated
the translocation dynamics to fruits. Therefore, both source organ (i.e.
leaves) and sink organ (e.g. roots, young shoots, developing seeds and
in this study in particular fruits) are important. Phloem mass flow
between these organs is generally believed to be driven by an
osmotically generated pressure gradient, a mechanism known as Münch’s
flow (Münch 1930; Nobel 2009). Hereby, photosynthates generated in
herbaceous crop leaves are mainly loaded into the phloem by active
transport (Hammond & White 2008). As the photosynthate concentration in
the phloem increases, its osmotic water potential decreases (more
negative). Consequently, water moves from the xylem into the phloem by
osmosis increasing the turgor pressure of the phloem (Lalonde et
al. 2003). This pressure pushes the phloem’s content down to the sink
organs where photosynthates are unloaded by active transport (Maynard &
Lucas 1982; Lalonde et al. 1999; Hölttä et al. 2006).
Hence, the photosynthate concentration of the phloem in sink organs is
decreased, which increases the overall water potential of the phloem.
Water molecules are hereby released from the phloem and return to the
xylem (De Schepper et al. 2013).
We investigated the effect of light intensity and application of Munch’s
theory to our study, while taking into account the positive correlation
between light intensities and translocation in leaves as described
earlier by Troughton et al. (1977) and Lanoue et al.(2018). We assumed an increasing amount of photosynthates to be loaded
into the phloem tissue under high light intensity compared to low light
intensity. This causes more water to flow into the phloem from xylem
resulting in a high turgor pressure of the phloem. Finally, a
high-pressure gradient is generated and photosynthate translocation is
promoted from sources to sinks. Summarised, by altering the light
intensity above the source leaf, we expected a linear increase in
relative photosynthate translocation into strawberry fruits. However, we
did not obtain a relationship between light intensity and relative
photosynthate translocation rate (Fig. 4B). This finding can be
attributed to the environment inside the exposure bag during PET
measurements. During each PET experiment, the controlled air was
supplied to the exposure bag at a rate of 400 mL
min-1. This flow rate was too low to renew the air
inside the exposure bag resulting in high relative humidity conditions
of 94.61 ± 0.97 %. This environment created small water droplets at
both the exposure bag and leaf surfaces, the former lowering the amount
of light reaching the leaf surface, and the latter inhibiting the gas
exchange through stomata (Ishibashi & Terashima 1995). This is shown by
the lower photosynthesis measured during the PET measurements (Fig. 4A;
grey dots) with respect to the light response curve of the same leaf.
Transpiration as a key effector on photosynthate
translocation to fruits
Transpiration rate was found to be the main factor affecting relative11C-photosynthate translocation to strawberry fruits.
This outcome is assumed to result from the decrease in xylem water
potential at the source leaf. When transpiration rate increases, leaf
xylem water potential becomes more negative (Alarcón et al. 2003;
Dodd et al. 2009; Steppe et al. 2015). This decrease in
water potential lowers water flow into the phloem from the xylem,
causing a decrease in phloem turgor pressure. Less turgor pressure
creates a decreasing pressure gradient between source and sink organs,
thus generating less pressure flow in the phloem (Sung & Krieg 1979).
This eventually results in a negative correlation between transpiration
rate and relative photosynthate translocation rate. In other words,
suppression of photosynthate translocation into fruits is caused by the
promotion of transpiration rate at the source leaf.
Because of the difference in translocation distance that depends on
plant size, the total amount of translocated11C-tracer to the fruits through the PET measurement
was smaller in large plants compared to small plants. However, there was
no difference in the relationship between the relative translocation
rate and transpiration rate for larger plants (“Red-27mm-L” and
“Red-18mm-L”) and smaller plants (i.e. “Red-27mm-S”,
“White-22mm-S” and “Green-18mm-S”). On the other hand, for the same
transpiration rate, a larger relative translocation rate is observed in
larger plants. Larger plants can hence maintain translocation of
photosynthates under higher transpiration rates (>c. 0.15 mmol H2O m-2s-1, i.e. the point from where zero photosynthate flow
is observed in small plants) than smaller plants. For the biggest fruits
on the large plants (i.e. “Red-27mm-L”) a less negative slope is
observed compared to all other fruits. This could be related to the
developmental stage of these fruits since they were already fully grown
(bright red colour) at the beginning of the experiment whereas the other
fruits, both red, white and green, were still growing. A linear increase
of the tracer concentration with time was observed for the last 40 min
of each TTC representing a constant flow of the tracer into the fruits.
Generally, the negative correlation between transpiration rate and
relative photosynthate translocation rate was similarly observed in
every PET measurement regardless of fruit’s developmental stage (colour
and size). Therefore, it is considered that suppression of photosynthate
translocation caused by the promotion of transpiration is a physical
phenomenon associated with xylem water potential. The physiological
factor related to fruit’s developmental stages, such as sink strength
(Marcelis 1996), did not affect the mechanism of photosynthate
translocation suppression associated with transpiration. This suggests
that, in protected horticulture, the transpiration management of source
leaf by environmental, for example vapour pressure deficit, control is
an effective method which can similarly control the photosynthate
translocation into fruits at all development stages. Furthermore, this
also suggests the possibility that the rate of photosynthate
translocation into fruits is promoted during night-time when
transpiration in a leaf is supressed.
Future potential of PET to investigate photosynthate
translocation
Positron emission tomography or PET was originally developed as a key
diagnostic tool used clinically to follow-up and treat diseases by
making use of positron-emitting radioisotopes. Given the in vivonature of this technique, its use has been extrapolated to plant science
to measure the transport of nutrients as well as phytohormones. Despite
the limited number of studies on plants, this imaging technique has
shown its applicability to investigate photosynthate translocation to
fruits (Kikuchi et al. 2008; Kawachi et al. 2011; Yamazakiet al. 2015; Hidaka et al. 2019). Nevertheless, the full
potential of 11C-positron emission tomography remains
largely unexploited (Hubeau & Steppe 2015). This can be related to the
fact that important aspects of whole-plant carbon allocation occur late
in development, i.e. when the plants are large. Most PET devices used in
research have not been adjusted to support experiments on bigger plants
which has restricted their use in plant biology (Karve et al.2015). Studies of large, full-grown plants are especially important
because much of our food is produced as seed late in development, i.e.
during the reproductive stage. In case of our study, it would have been
advantageous if, aside from the fruits, also the source leaf and other
leaves could have been imaged at the same time. As such it could be
determined which fraction of the assimilated11CO2-tracer is used for storage,
respired to the atmosphere or ends up in the fruits/other leaves. These
difficulties could be overcome by making use of clinical PET, which is
developed for human imaging, as these systems have two main advantages.
Firstly, these imaging devices allow visualisation of bigger objects
since they are characterised by a transverse and axial field of view
(FOV) up to 85 and 26 cm, respectively (Vandenberghe & Marsden 2015;
Vandenberghe et al. 2016). This is generally larger than the ones
of small animal PET scanners (transverse and axial FOV of 10 and 7.5 cm
in this study). Besides, clinical PET scanners are equipped with a
moving bed on which the plant can be placed which enables the
visualisation of bigger plants. A drawback of clinical PET systems is
the lower spatial resolution (c. > 5 mm
(Vandenberghe et al. 2016)) compared to the sub-millimetre
resolution of small animal PET scanners (España et al. 2014; Fineet al. 2014). Hence, visualisation of smaller sized plant
structures (e.g. petioles and peduncles) will be complicated using
clinical PET systems in contrast to bigger sized fruits and whole
leaves. The second advantage of the clinical PET systems is the fact
that they are nearly exclusively used in combination with structural
imaging like computed tomography (CT) or magnetic resonance imaging
(MRI). Consequently, the functional information about the radiotracer
under study provide by PET is reinforced with anatomical data provided
by CT or MRI. Despite the intensive occupancy of these clinical PET
systems, we believe that studies making use of these imaging devices
will make a major contribution to reveal complicated in vivointeractions in plants (e.g. link between xylem and phloem tissue
in larger trees).
Conclusion
Non-invasive, real time 11C-photosynthate
translocation from source leaf to strawberry fruits was successfully
dynamically visualised by making use of PET. To our understanding this
study is the first to report dynamic tracing of photoassimilates in 3D.
By manipulating the light intensity at the source leaf, we expected to
affect the translocation rate. However, we found that photosynthesis is
not the main driver for the photosynthate translocation to fruits since
our results suggest transpiration being the key effector. This is an
important finding with regard to the optimisation of commercial
strawberry production. Furthermore, we acknowledge the advantage ofin vivo experiments based on radiotracers and suggest that plant
sciences could benefit from using clinical PET devices to study intact
plants.