INTRODUCTION
Photosynthesis, as measured by gas exchange, is typically assessed by
the three canonical biochemical limitations of photosynthesis: the
rubisco limitation, where carbon dioxide uptake is modeled assuming
ribulose 1,5-bisphosphate (RuBP)-saturated rubisco kinetics; the RuBP
regeneration limitation, where carbon dioxide uptake is modeled assuming
a fixed rate of RuBP use as allowed by the production of electron
transport products, ATP and NADPH; and the triose phosphate utilization
(TPU) limitation, where carbon dioxide uptake is modeled as the rate of
production of end products, freeing inorganic phosphate from organic
phosphates (McClain & Sharkey 2019). The TPU limitation is unique among
the three biochemical limitations in that it is limited by downstream
processes, rather than just at rubisco. The dislocation of the
limitation from rubisco means that regulatory mechanisms are engaged to
slow down the rate of carbon assimilation (A ) so as not to
outpace the rate of end-product synthesis. Energy-dependent quenching
(qE ) is activated (Sharkey, Berry &
Sage 1988) by elevated ΔpH across the thylakoid membrane, one component
of proton-motive force (PMF) (Kramer & Crofts 1996). The elevated ΔpH
results from kinetic and thermodynamic restrictions on the ATPase due to
lowered levels of available inorganic phosphate (Sharkey & Vanderveer
1989). In addition, rubisco activation state decreases (Sharkey, Seemann
& Berry 1986a; Socias, Medrano & Sharkey 1993), which may alleviate
pressure on phosphate pools by limiting the maximum rate that carbon can
be added to the organic phosphate pool. Because TPU limitation restricts
the rate of photosynthesis rather than the availability of light, there
is a potential for photodamage unless regulatory mechanisms are engaged
(Powles 1984; Pammenter, Loreto & Sharkey 1993; Li, Müller-Moulé,
Gilmore & Niyogi 2002).
These regulatory mechanisms are the only aspects of TPU limitation
typically observed in steady-state gas exchange. While TPU limitation
results in, and can be assessed through, gas exchange as
O2- and CO2-insensitive photosynthesis
(Sharkey 1985) or reverse sensitivity to O2 (Viil,
Laisk, Oja & Pärnik 1977) or CO2 (Jolliffe & Tregunna
1973), it is easier to assess by the decline in electron transport rate
associated with qE when
CO2 is increased or O2 is decreased. The
appearance of transient effects on photosynthesis associated with TPU
limitation (Ogawa 1982; Walker, Sivak, Prinsley & Cheesbrough 1983)
lead us to believe that, in the steady state, the rate of photosynthesis
is not set by TPU, but instead, the rate is set by regulatory mechanisms
that match the rates of carbon input to and carbon output from the
organic phosphate pool.
The nitrogen required for rubisco and photosynthetic electron transport
far exceed those required for TPU and subsequent end product synthesis
(Evans & Clarke 2019). When TPU occurs, rubisco is deactivated andqE is increased reducing the efficiency of
nitrogen use in both carbon metabolism and electron transport. Because
TPU capacity is relatively cheap and entering TPU limitation forces
deactivation of systems which use much more nitrogen, an ideal plant
would never experience TPU limitation under physiological conditions.
However, TPU limitation is commonly seen when the photosynthetic rate is
only a few percent higher than what the plant experiences in ambient
conditions (Yang, Preiser, Li, Weise & Sharkey 2016). There are a few
possible reasons why excess TPU capacity would be detrimental. A precise
balance of phosphate flux could control stromal inorganic phosphate
concentration, affecting the partitioning of carbon into starch (Preiss
1982; Escobar-Gutiérrez & Gaudillère 1997). If TPU capacity were in
excess, it could also limit the ability to build up a PMF across the
thylakoid membrane because there would be plentiful phosphate available
to the ATPase, preventing any kinetic or thermodynamic restriction to
proton flow. The elevated ΔpH and consequent low luminal pH can activate
energy-dependent quenching mechanisms that dissipate light energy to
safeguard the photosystems.
If TPU capacity is inexpensive in terms of nitrogen cost, but is
typically just above ambient photosynthetic rates, we would expect that
TPU capacity is plastic. It has been found that TPU capacity is
flexible, and in many cases changes in response to environmental
conditions. Plants grown at low temperature can develop additional
sucrose synthesis enzymes (Cornic & Louason 1980; Guy, Huber & Huber
1992; Holaday, Martindale, Alred, Brooks & Leegood 1992) which
alleviates cold-induced TPU limitation (Sage & Sharkey 1987). Plants
with reduced access to CO2 have reduced TPU capacity to
match their lowered photosynthetic rate (von Caemmerer & Farquhar 1984;
Sharkey & Vassey 1989). It has therefore been shown that TPU capacity
can both increase and decrease in response to environmental conditions.
This is reflected in environmental surveys, and plants have rarely been
found to be TPU limited under ambient conditions in the field (Sage &
Sharkey 1987; Ellsworth, Crous, Lambers & Cooke 2015). For this reason,
TPU limitation is often not included in global models of photosynthesis
(Lombardozzi et al. 2018; Rogers et al. 2020).
Ideally, if a plant is TPU limited, it will increase its TPU capacity to
maximize the overall rate of photosynthesis, but it is also possible
that rubisco capacity and electron transport capacity will be decreased
to match TPU capacity. We tested the acclimation of plants to TPU
limitation by exposure to elevated CO2 to determine
whether plants eventually stop being TPU limited, and if they achieve
this by increasing their TPU capacity. In addition, we established a
timeline of the regulatory features surrounding TPU limitation, from how
the plant handles the initial influx of energy until the plant engages
slower regulatory features, such as rubisco deactivation and
energy-dependent quenching.