Temperature affects the development, growth, and physiological processes of plants (Asseng et al., 2013). The air temperature of the earth experienced significant increases during the 1950s–2000s (Intergovernmental Panel on Climate Change [IPCC], 2007). Global warming shows variable diurnal and seasonal warming patterns known as asymmetric warming. Increases of temperature amplitude in winter and spring are greater than in autumn and summer. Also, warming is greater at night than during the day, which describes a phenomenon commonly referred to as asymmetric warming (Intergovernmental Panel on Climate Change [IPCC], 1998; Su et al., 2015). These climatic trends are likely to further increase until 2050 (Chavas et al., 2009). Winter wheat is normally cultivated over the winter and spring seasons (Liu et al., 2014). As a warm- and cool-season crop, this sort of climate warming is likely to profoundly influence winter wheat. However, experiments simulating the impact of warming on winter wheat are mostly based on mean air temperature, without consideration for diurnal warming asymmetry.

Warming was reported to have dramatic effects on the morpho-physiological processes of plants, including altered crop development and growth (Tao and Zhang, 2013), diminished functionality of photosystem II (PSII) (Atkin and Tjoelker, 2003), and alleviated antioxidant systems (Martinez et al., 2014), and to influence the duration of grain filling (Liu Z. et al., 2013). An increase in night temperature by 1.1°C promotes grain-filling rates both in superior and inferior grains, ultimately resulting in a significantly increased 1,000-grain weight in wheat (Chen et al., 2014). In a 5-year experiment, it is found that the grain yield of winter wheat increased by 16.3%, and the grain-filling rate of inferior grain was stimulated by a full-day temperature increase of 1.5°C (Tian et al., 2012). Plant hormones are key regulators of grain development under different plant growth conditions (Yang et al., 2006). Morris et al. (1993) observed a large transient rise in cytokinin (CTK) levels after anthesis in the developing grains of wheat and rice that significantly affected the duration of grain filling and grain size. Lur and Setter (1993) reported that indole-3-acetic acid (IAA) content rapidly increased in maize (Zea mays) kernels at nearly 10 days after anthesis, which resulted in an increase in assimilate transport to the developing kernels and ultimately resulted in higher kernel weights.

Crop yield is closely related to net photosynthetic assimilation process during the grain-filling period in wheat. The photosynthesis of the flag leaf contributes the most to final grain dry weight in winter wheat. Study of the flag leaf in response to climate warming that occurs during the winter and spring seasons is crucial. Photosynthesis is the most temperature-sensitive process in plants. Some species can adapt to temperature change, as indicated by shifts in the thermal optimum and improved photosynthetic rates at new growth temperatures (Chen et al., 2014). Martinez et al. (2014) found that when temperature increases to the optimal range of the plant, the photosynthetic rate is expected to increase. However, the mechanism of temperature acclimation regarding photosynthesis is still unclear. Some studies assumed that this acclimation is primarily due to the increased regeneration capability of ribulose-1,5-bisphosphate (RuBP) and the increased carboxylation capacity of Rubisco (Wang et al., 2011). According to the models of Farquhar et al. (1980), the balance between RuBP regeneration and carboxylation, as indicated by the maximum rate of carboxylation (V_cmax) versus the maximum rate of photosynthetic electron transport (J), determines the temperature dependence of photosynthesis. However, evidence from previous experiments does not clarify the mechanism of photosynthetic change under climate warming during the entire life cycle of winter wheat.

Photosynthetic properties and senescence in plants are inherently programmed, but both are influenced largely by environmental conditions (Riikonen et al., 2009). Lobell et al. (2012) suggested that chlorophyll content could determine the photosynthetic capacity of plants, which has a positive correlation with the chloroplast membrane stability, and could be used to screen for heat stress in winter wheat. Reactive oxygen species (ROS), which are involved in the physiological metabolism and life processes of plants, inhibit photosynthesis. To prevent ROS accumulation, plants have developed certain antioxidant systems. The effects of heat stress on membrane peroxidation, reduction of PSII activity, and chlorophyll loss are caused by oxidative stress related to overproduction of ROS (Djanaguiraman et al., 2010). Most of the studies on both thermal acclimation and the characteristics of oxidative metabolism have been conducted using constant temperature regimes and have occurred in greenhouse or growth chambers (Atkin and Tjoelker, 2003; Xu et al., 2008). However, how antioxidant capacities and the production of ROS respond to asymmetric warming conditions under field conditions remains to be determined.

It has been shown that abiotic stress priming or adaptation during vegetative periods can improve plant stress tolerance during subsequent reproductive growth stages (Cui et al., 2015). For example, winter wheat subcellular antioxidant systems were activated under cold priming, and consequently, the tolerance to subsequent cold stress was enhanced (Li et al., 2014a). Pre-anthesis high-temperature priming enhanced plant photosynthesis due to the expression of the photosynthesis-responsive gene RcaB (encoding Rubisco activase B) and the major chlorophyll a/b-binding protein gene Cab (Wang et al., 2011). Li et al. (2014b) found that the expression of antioxidant enzyme-related genes such as Cu/Zn-SOD, which encodes chloroplast Cu/Zn superoxide dismutase, and Mn-SOD, which encodes mitochondrial manganese superoxide dismutase, was upregulated under drought acclimation. Heat priming using a moderate temperature can improve tolerance to subsequent heat stress in plants (Larkindale and Huang, 2004). However, it is still unclear if asymmetrical warming during vegetative periods can improve the post-anthesis physiological activities of winter wheat.

We have reported that winter night-warming promoted pre-anthesis plant growth and improved post-anthesis net photosynthetic rates, ultimately increasing grain yield (Fan et al., 2015). In the present study, winter wheat plants were exposed to night-warming during the winter and spring seasons to examine grain-filling processes, grain sucrose content, grain endogenous hormones, photosynthetic capacity, antioxidant capacity, the expression of photosynthesis-responsive genes, and antioxidant enzyme-related genes during the reproductive growth period. Our main objective was to determine whether winter and spring night-warming could improve the post-anthesis source activity and sink strength of winter wheat and to clarify the possible physiological mechanisms involved. We hope the results will contribute to better understanding of the physiological mechanisms causing wheat yield increase under winter and spring night-warming.

The objective of this study was to test the hypothesis that winter and spring night-warming upregulate the source and sink activities of winter wheat, which would be beneficial for grain yield formation. We observed that warming stimulated sink activity, which could therefore accelerate source carbohydrate depletion and, in turn, advance source activity (Tian et al., 2012). Sink formation (e.g., grain weight and grain number) and sink activity (e.g., grain-filling rate) enhance source productivity (e.g., photosynthesis) (Chen et al., 2014). Our results demonstrated that WW increased flag leaf area (Figure 2) and slightly increased specific leaf weight during the grain-filling stage (Figures 2E,F). In our previous study, we found that from the sowing to jointing stages and from the jointing to anthesis stages the wheat plants under WW and SW treatments showed higher growth rates (Fan et al., 2015). The mean night temperatures of the WW and ST treatment increased by 1.56–1.67°C and 1.78–1.92°C, respectively. Chen et al. (2014) indicated that increased night temperature can increase cell metabolism, the nighttime respiration rate of leaves and the growth rate of leaves. Under night-warming conditions, the number of leaf cells and plant metabolism become more vigorous. This may be the reason for the post-anthesis source improvement by night-warming, and this phenomenon benefited crop canopy light interception and contributed to higher grain weight.

Our previous study also showed that the growing degree days of the vegetative growth stage (from sowing to anthesis stage) increased by winter night-warming; thus, the pre-anthesis period was reduced (Fan et al., 2015). Winter wheat in the Yangtze River Basin of China, where the experiments were conducted, usually experiences extreme high-temperature weather during the grain-filling stage (Cao et al., 2011). The shortened pre-anthesis period caused by WW and SW may shift wheat plants in the post-anthesis period forward into a relatively cooler environment than the control (Liu et al., 2010). In addition, both WW and SW treatments increased the post-anthesis period of winter wheat, increasing the grain-filling time. Thus, wheat plants under WW and SW treatments benefited from complete physiological processes and grain-filling procedures. In the present study, WW significantly increased the post-anthesis flag leaf Pn and the maximum A_sat of Yangmai-13 and significantly increased the V_cmax of both cultivars (Table 1). Furthermore, the J of Yangmai-13 was increased by WW and SW (Table 1). Yamori et al. (2005) demonstrated that V_cmax is known to be enhanced with high-temperatures (the day/night air temperatures were 30/25°C for 1.5 months), while J could limit the photosynthesis rate under high-temperatures. Under warming treatments, the enhanced leaf PSII performance was due to the increased activities of photosynthesis-related enzymes and the enlargement of the leaf area (Martinez et al., 2014). These results indicate that moderate warming during the winter and spring improved the flag leaf photosynthetic capacity and Rubisco activity of winter wheat. Chlorophyll content during the grain-filling stage has been used as an efficient indicator of photosynthetic capacity in wheat cultivars (Christ and Hörtensteiner, 2014). Our study showed that WW and SW increased the chlorophyll content of flag leaves during the early stage of grain filling for both cultivars, while WW increased this chlorophyll content more than SW did (Figure 3). This finding indicates that winter night-warming enhanced photosynthesis in flag leaves by enhancing Rubisco carboxylation capacity, electron transport efficiency, ATP synthesis, and Calvin cycle processes. Consequently, our findings suggest that night-warming during winter could improve flag leaf morphology and photosynthetic capacities, which favor the grain-filling process of winter wheat.

Improved efficiency of photosystems also prevents the generation of ROS (Gallé et al., 2007). Wang et al. (2011) reported that pre-anthesis heat priming increases grain yield against subsequent high-temperatures during the grain-filling stage, and their finding was attributed to the improved photosynthetic and antioxidative activity in the acclimated plants. Our study showed that the WW treatment reduced post-anthesis ROS production (Figures 4A–D). This result indicates that plants under winter night-warming maintained a sufficient antioxidant capacity to avoid ROS accumulation. MDA content is used to evaluate the redox and osmotic adjustment status, which is important in the adaptation of plants to environmental stresses (Hung and Kao, 2004). Our study showed that the MDA content of flag leaves was lower for WW and SW than for NW for both cultivars after anthesis (Figures 4E,F). This indicates that WW and SW increased the capacity of the antioxidant system for scavenging ROS and regulating the accumulation of ROS. Similar results have been reported for Kobresia pygmaea (Yang et al., 2012) and Betula pendula (Riikonen et al., 2009). In addition, our study showed that WW and SW increased the SOD, POD, and CAT activity of flag leaves during the early grain-filling stage for both cultivars (Figure 5). This result suggests that the observed increase in SOD and POD activity would compensate for ROS accumulation. As CAT breaks down H₂O₂, an increase in CAT activity would have resulted in a reduction in H₂O₂, thereby attenuating oxidative damage of membrane. These data suggest that winter and spring night-warming increase post-anthesis antioxidant capacities to attenuate oxidative membrane damage, which is beneficial for improving flag leaf physiological activities.

To further illustrate the underlying mechanisms of increased post-anthesis photosynthesis and antioxidant properties induced by winter and spring night-warming, we analyzed the expression of photosynthesis-responsive and antioxidant enzyme-related genes. Wang et al. (2014) reported that Rubisco activase is a major limitation to photosynthetic capacity and a potential target for genetic manipulation that can enhance wheat productivity under high-temperatures. Kurek et al. (2007) reported that photosynthesis reduction is correlated with Rubisco deactivation resulted in Rubisco activase inhibition under high-temperature. In our study, RcaB expression in flag leaves was upregulated by WW (Figure 6A), and such a trend was associated with the enhanced photosynthetic capacity of flag leaves in WW plants compared with NW plants. Rampino et al. (2006) indicated that Cab was among the genes encoding a major chlorophyll a/b-binding polypeptide in the light-harvesting complex. Our study showed that the effects of WW and SW on Cab expression were not significant, except for Yangmai-13 under SW (Figure 6B). Furthermore, WW upregulated the expression of the antioxidant enzyme-related genes Cu/Zn-SOD (in Yangmai-13), Mn-SOD (in Yannong-19), and CAT (in Yangmai-13) compared with NW (Figures 6C-E). These higher antioxidant activities in the WW plants were associated with the upregulated expression of the antioxidant enzyme-related genes. The above evidence suggests that moderate night-warming during winter increases the expression of both photosynthesis-responsive and antioxidant enzyme-related genes, which directly regulate higher leaf photosynthesis and oxidation resistance abilities during the early grain-filling stage.

Our previous results showed that winter and spring night-warming increases the grain weight of both cultivars (Fan et al., 2015). Furthermore, we found similar positive effects of night-warming on the grain-filling rates during the early filling stage for both cultivars (Table 2), which suggests that winter and spring night-warming would benefit the grain filling of winter wheat. In our experiment, we used a moderate warming temperature of 1–2°C during the night, which is in line with the global warming rate projected by the Intergovernmental Panel on Climate Change (IPCC), resulting in the advancement of the phenophases of anthesis and maturity (Fan et al., 2015). This could shift the post-anthesis phase of winter wheat to more optimal temperature conditions for leaf photosynthesis and grain filling (Tian et al., 2012).

Our study showed that the grain sucrose content increased under WW (Figure 7A) and that the grain SS activity increased significantly under WW and SW for both cultivars (Figure 7B). Grain filling is mainly a process of starch biosynthesis and accumulation, and sucrose is the main substrate for the grain-filling process. SS activity is positively related to dry matter accumulation during grain filling, and SS activity is believed to be important in determining sink strength. Our study indicated that night-warming enhances sink strength and increases the amount of available sucrose for grain filling. CTK and IAA play important parts in regulating grain filling in cereals; the CTK content in rice spikelets is significantly positively correlated with grain development (Hao et al., 2009; Liu Y. et al., 2013). Xu et al. (2007) reported CTK and IAA regulate endosperm cell division in developing rice seeds. High IAA levels in a sink organ can create “attractive power,” leading to increased CTK levels in grains (Liu Y. et al., 2013). Our study showed that WW significantly increased the grain IAA and CTK concentrations at 14 DAA for both cultivars (Figures 7C,D). The present results indicate that IAA and CTK regulate grain filling of winter wheat in the early grain-filling stage, possibly due to the manipulation of endosperm cell division, thus creating sink strength.

Our study also showed that the increase in sink capacity was higher in Yangmai-13 (vernal type) than in Yannong-19 (semi-winter type) under WW. The optimal temperature during the vernalization phase is higher for the vernal type than for the semi-winter type of winter wheat (Singh et al., 2013). The warmer conditions under WW were more adaptive for Yangmai-13 growth and were particularly beneficial for spikelet and floret differentiation, which occurred during winter. We have previously reported that winter night-warming improves the pre-anthesis crop growth rate and enlarges the post-anthesis source of winter wheat, which is beneficial for grain dry matter accumulation during the grain-filling stage (Fan et al., 2015). Prolonged grain-filling duration certainly benefits grain formation. Consequently, the above evidence suggests that moderate nigh-warming during winter and spring would be beneficial to accelerate the grain-filling rate and enhance sink strength during the early grain-filling stage of winter wheat.

We concluded from the results of the study that night-warming promoted source productivity by enlarging the flag leaf area and improving photosynthetic capacity. Night-warming also increased the antioxidant capacities, which attenuated oxidative membrane damage. This was seemingly controlled at the transcription level as indicated by the changes in the expression of both photosynthesis-responsive and antioxidant enzyme-related genes. Night-warming enhanced sink strength and accelerated the grain-filling rate; however, the increases were greater during winter night-warming than during spring night-warming. The above evidence demonstrates that winter night-warming improves grain filling and the flag leaf physiological characteristics of winter wheat, resulting in an improved performance of wheat crop.

YF, TD, ZH, and ZT designed the experiment. YF conducted the study, collected and analyzed the data, and prepared the draft. YY and CH helped in sampling and the measurements of physiological parameters. DJ and CM helped in drafting the manuscript and the interpretation of the results. MA helped in revising the manuscript and the responses to the comments.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.