**1. Introduction**

Stomata, from the Greek word "stoma" meaning mouth, are small pores that distributed on the epidermis of plant leaves. Their structures consist of two guard cells around a pore. For optimum efficiency, stomata must balance the gas exchange between inside and outside the leaf, in order

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tomaximize CO2uptake forphotosynthetic carbonassimilation(*P*N) andtominimize waterloss through transpiration (*E*). Although the cumulative area of stomatal pores only represents a small fraction of the leaf surface, typically less than 3%, the CO2 uptake and water loss pass through these pores. When fully open, they can promote water evaporation equivalent to onehalf of a wet surface of the same area [1]. To cope with environmental stress during growth condition, plants must adjust and regulate the stomatal opening/closing process to obtain optimized transpiration and leaf water status.

On the other hand, studying the evolutionary adaptation and natural variation of stomatarelated genes may represent an essential step for better understanding the mechanisms involved in the stomatal adjustment and regulation. In fact, stomata have probably undergone a crucial adaptation occurring 400 million years ago, it enabled plants to thrive on land. To survive in the dry atmosphere, plants must maintain a reasonable level of gas exchange necessary for *P*N and *E*, in order to protect against desiccation [2]. In addition, the natural variation in stomata-related genes across different cultivars (from different origins) of particular species may indicate differing selection pressures allowing better adaptation against environmental stress [3].

To get a deeper understanding, the study of the relationship between genotype and phenotype at the organism–environment interface by identifying traits that respond to differing environ‐ mental pressures and uncovering the genetic basis for variability in these traits is highly requested. Recent researches have shown that the mode of action of stomatal movement depends on the combination of environmental and intracellular signals. These external factors (e.g., CO2, biotic and abiotic stresses, and additionally different plant hormones) and internal signals (e.g., ion exchange, metabolites, catalyze of enzyme, and gene structure or expression) simultaneously affect stomatal dynamics, forming a complex framework behind acclimation responses of plants under fluctuating and stressed environments. The empirical evidences related to stomatal dynamics provide strong promotion for the development of model stimulating stomatal dynamics, which remains difficult to achieve so far. In this chapter, we aim to give a multidimensional review about recent works describing multiple environmental and internal factors, such as elevated CO2, heat stress, light fluctuations, ion channel, and stomata-related genes [4–8]. Furthermore, we discussed expended research perspective regarding stomatal evolution, natural variations of stomatal traits, interactions with life history, and theoretical modeling.

## **2. External environments**

#### **2.1. Interactive effects of elevated CO2 and heat wave**

The global change, leading to frequent occurrence of atmospheric CO2 enrichment and heat wave, inevitably affects the development and final productivity of plants. Most climate impact studies rely on changes in means of meteorological variables, such as temperature and rainfall, to estimate the potential climate impacts on agricultural production. However, extreme meteorological events, e.g., a short period of abnormally high temperatures, can have a significant profound and lasting effect on canopy transpiration, crop growth, and final yield [9].

During heat stress, elevated CO2 has probably less effect on C3 plants as compared to C4 plants [10]. In fact, elevated CO2 can increase water-use efficiency (WUE) by decreasing stomatal conductance (*g*s) and *E* [11], which may increase tolerance to acute heat. It was shown that the reduction in *g*s (stomatal opening) is about 20% for C3 and 50% for C4 species [10, 12, 13]. The lower *g*s in C4 plants may induce lower transpiration (water loss) and thus higher leaf tem‐ peratures, which may increase heat-related damage in C4 plants as compared to C3 plants in the same habitat.

Since evaporative cooling is essential to avoid heat damage in leaves exposed to full sunlight, and time scales of stomatal adjustments are longer than fluctuations in solar irradiance within a canopy, the question arises whether elevated CO2 can mitigate damage over transpiring leaves from extreme high temperature by decreasing *g*s. If this is the case, then adaptation for cooling would appear as a more imperative driver for stomatal adjustments than the potential increase in carbon gain. To test this hypothesis, intact leaves of maize were subjected to a substantial reduction *P*N due to 45°C heat stress cycle for 1hour[14]. Our previous finding showed that elevated CO2, either during plant growth or co-heat period, does not improve the foliar thermotolerance against heat stress (Figure 1). With the lower *P*<sup>N</sup> and higher *g*<sup>s</sup> and subcellular CO2 pressure (*C*<sup>i</sup> ) following the acute heat stress treatment, a non-stomatal inhibition of *g*<sup>s</sup> occurs, contrary to other studies showing a stomatal adjustments in response to high temperature stress in grape leaves [15, 16]. In the meantime, the sudden reversal of stomatal responses to leaf temperature and CO2 between 40°C and 45°C (Figure 1) suggests that to avoid damage, plants enhance the stomatal opening, leading to an increase in evapo‐ rative cooling.

Some studies compared elevated CO2 effects with tolerance to heat stress in relatively heatsensitive vs. heat-tolerant species or in species with different photosynthetic pathways [4, 17– 20]. As an example, two corn cultivars (B73 and B106) were previously reported as contrasting heat stress tolerance from field investigation and evaluations [21]. When comparing the effects of elevated CO2 and heat stress from field-based investigation using these corn cultivars, our previous results showed a reversible response of two cultivars regarding to photosynthetic activity (Figure 2), which might be due to intricate reasons: 1) change in physical function of stomatal regulation by decreasing transpiration and optimized water conservation at intact leaves scales; 2) change in kinetic activities of photosynthetic regulatory enzymes, i.e., rubisco (ribulose 1,5-bisphosphate carboxylase/oxygenase), PEPase, and MDHase (Table 1), which agrees with some reports [22, 23]; and 3) disorder of metabolite flux in Calvin cycle due to heat stress.

#### **2.2 Fluctuating light effects**

tomaximize CO2uptake forphotosynthetic carbonassimilation(*P*N) andtominimize waterloss through transpiration (*E*). Although the cumulative area of stomatal pores only represents a small fraction of the leaf surface, typically less than 3%, the CO2 uptake and water loss pass through these pores. When fully open, they can promote water evaporation equivalent to onehalf of a wet surface of the same area [1]. To cope with environmental stress during growth condition, plants must adjust and regulate the stomatal opening/closing process to obtain

On the other hand, studying the evolutionary adaptation and natural variation of stomatarelated genes may represent an essential step for better understanding the mechanisms involved in the stomatal adjustment and regulation. In fact, stomata have probably undergone a crucial adaptation occurring 400 million years ago, it enabled plants to thrive on land. To survive in the dry atmosphere, plants must maintain a reasonable level of gas exchange necessary for *P*N and *E*, in order to protect against desiccation [2]. In addition, the natural variation in stomata-related genes across different cultivars (from different origins) of particular species may indicate differing selection pressures allowing better adaptation against

To get a deeper understanding, the study of the relationship between genotype and phenotype at the organism–environment interface by identifying traits that respond to differing environ‐ mental pressures and uncovering the genetic basis for variability in these traits is highly requested. Recent researches have shown that the mode of action of stomatal movement depends on the combination of environmental and intracellular signals. These external factors (e.g., CO2, biotic and abiotic stresses, and additionally different plant hormones) and internal signals (e.g., ion exchange, metabolites, catalyze of enzyme, and gene structure or expression) simultaneously affect stomatal dynamics, forming a complex framework behind acclimation responses of plants under fluctuating and stressed environments. The empirical evidences related to stomatal dynamics provide strong promotion for the development of model stimulating stomatal dynamics, which remains difficult to achieve so far. In this chapter, we aim to give a multidimensional review about recent works describing multiple environmental and internal factors, such as elevated CO2, heat stress, light fluctuations, ion channel, and stomata-related genes [4–8]. Furthermore, we discussed expended research perspective regarding stomatal evolution, natural variations of stomatal traits, interactions with life

The global change, leading to frequent occurrence of atmospheric CO2 enrichment and heat wave, inevitably affects the development and final productivity of plants. Most climate impact studies rely on changes in means of meteorological variables, such as temperature and rainfall, to estimate the potential climate impacts on agricultural production. However, extreme meteorological events, e.g., a short period of abnormally high temperatures, can have a

optimized transpiration and leaf water status.

environmental stress [3].

76 Applied Photosynthesis - New Progress

history, and theoretical modeling.

**2. External environments**

**2.1. Interactive effects of elevated CO2 and heat wave**

Leaves are always subjected to rapidly fluctuating irradiance due to motion of sunflecks and clouds that may span two orders of magnitude from light compensation points of shadeadapted leaves to almost full irradiance intensities [25]. Such environmental fluctuations occur at second scales, which is much shorter than the time needed for stomatal adjustments (2–60

**Figure 1.** Dynamic changes of photosynthetic parameters during acute heat stress cycles. Lines with same color stand for treatment at same exposure CO2 concentrations. Symbols GambEamb, GambEelv, GelvEamb, and GelvEelv represent grown and exposed at ambient [CO2], grown at ambient [CO2] but exposed at elevated [CO2], grown at elevated [CO2] but exposed at ambient [CO2], and both of grown and exposed at elevated [CO2]. *Vertical bars* represent S.E. for *n* = 9 (see [14]).

min.) [26]. For leaves with slowly adjusting stomata, rapid fluctuations at shorter time scales could push leaf hydraulic and thermal status beyond operational limits resulting in xylem cavitation, overheating, or wilting.

Although the phenomena underlying dynamic responses of photosynthesis to sunflecks (such as induction requirements) were studied by physiologists and biochemists earlier [26], their role in sunfleck utilization was not recognized until the early 1980s. Evidence for the light activation requirement of the primary carboxylating enzyme, Rubisco, was first uncovered in the 1960s [27]. The components underlying induction, especially stomatal behavior, are complex and are dependent on environmental and developmental factors as well transient light changes. It was reported that water stress could reduce *g*s in shade-grown, but not in sun-

The physiology and genetics of stomatal adjustment under fluctuating and stressed environments http://dx.doi.org/10.5772/62223 79

**Figure 2.** Heat induced decrease of photosynthesis and stomatal conductances in B76 and B106. Black and grid bars represent ambient and elevated [CO2], respectively. Ratio of photosynthesis and stomatal conductances under heat stress over control in B76 and B106 was shown in inserted panel. (Qu et al. 2016, unpublished data).

grown for the leaves of a *Populus* species; drought also could lead to faster induction gain in shade-grown, but not in sun-grown for the leaves during simulated sunflecks [28] .

min.) [26]. For leaves with slowly adjusting stomata, rapid fluctuations at shorter time scales could push leaf hydraulic and thermal status beyond operational limits resulting in xylem

Leaf Temperature [°C]

**Figure 1.** Dynamic changes of photosynthetic parameters during acute heat stress cycles. Lines with same color stand for treatment at same exposure CO2 concentrations. Symbols GambEamb, GambEelv, GelvEamb, and GelvEelv represent grown and exposed at ambient [CO2], grown at ambient [CO2] but exposed at elevated [CO2], grown at elevated [CO2] but exposed at ambient [CO2], and both of grown and exposed at elevated [CO2]. *Vertical bars* represent S.E. for *n* = 9 (see

40 45 45 40 35 35 35

40 45 45 40

WUE [µmol mmol–1

 KPa]

5

6

VPD [KPa]

7

*gs* [mol mol–2 s–1]

**D**

**B**

GambEamb GambEelv GelvEamb GelvEelv

**E F**

Although the phenomena underlying dynamic responses of photosynthesis to sunflecks (such as induction requirements) were studied by physiologists and biochemists earlier [26], their role in sunfleck utilization was not recognized until the early 1980s. Evidence for the light activation requirement of the primary carboxylating enzyme, Rubisco, was first uncovered in the 1960s [27]. The components underlying induction, especially stomatal behavior, are complex and are dependent on environmental and developmental factors as well transient light changes. It was reported that water stress could reduce *g*s in shade-grown, but not in sun-

cavitation, overheating, or wilting.

35

*E* [mmol m–2 s–1]

[14]).

200

300

*Ci* [µmol mol–1]

*PN* [µmol m–2 s–1]

400

**A**

78 Applied Photosynthesis - New Progress

**C**

In the naturally fluctuating environment, the temporal disconnect between *g*s and *P*<sup>N</sup> means the coordination between carbon gain and water loss (and, therefore, WUE) is far from optimal ([29]; Figure 3). Photosynthetic induction state is a complex function of light-dependent stomatal opening and closing responses and the time courses of light-regulated enzyme activation and deactivation. All these combined factors determine the potential light-saturated *P*<sup>N</sup> at any given time and therefore the potential *P*<sup>N</sup> that can be achieved during a fluctuating light (shade-fleck). Under this condition, responses of *g*s and *P*<sup>N</sup> are not always synchronized, as stomatal movements can be an order of magnitude slower than the more rapid photosyn‐ thetic response to the same environmental stimuli ([30, 31]; Figure 3).


**Table 1.** Enzyme activities of PEPC, NADP-ME, and NADP-MDH for B76 vs. B106 grown ambient and elevated [CO2]. Values of control experiments were shown in brackets (Qu et al. 2016, unpublished data).

**Figure 3.** Photosynthesis and stomatal conductance in response to naturally light regime (Qu et al. 2016, unpublished data).
