Soybean (Glycine max) is one of the most important food crops. One of the major compositions in soybean seeds is protein, which is necessary for human nutrition; soy is also a valuable source of edible oil (Medic et al., 2014). During the developmental stage, soybeans are often at risk of high temperature stress. The Intergovernmental Panel on Climate Change (IPCC) reports that future climate change will expose crops to heightened average temperatures and increase the frequency of short-term high temperatures (Solomon, 2008). High temperature causes significant agricultural losses, affects nutrient cycling in many crops, and leads to substantial yield decline (Asseng et al., 2014, Schauberger et al., 2017; Zhao et al., 2017). When soybeans suffer high temperatures during the seed filling stage, their yields are reduced and their seed compositions changed (Medic et al., 2014; Song et al., 2016; Nakagawa et al., 2020). For legumes such as faba beans, various genetic approaches have been used to breed for heat stress tolerance varieties (Lavania et al., 2015; Lavania et al., 2016). Global climate model projections indicate that nighttime warming rises more rapidly than daytime warming and is the primary driver of global warming (Davy et al., 2016), reducing diurnal temperature differences (Vose et al., 2005). A study by our research group found that the diurnal temperature difference had a greater effect on soybean protein and oil contents relative to the average daily temperature, suggesting that night temperature plays an important role in soybean protein and oil accumulation (Song et al., 2016). Therefore, studying the effects of nighttime warming on soybean growth and yield will provide a theoretical basis for developing future food security strategies in China.

Several studies have confirmed the effects of temperature on soybean seed growth in different developmental stages, duration of high temperature treatments, and intensities of temperatures (Gibson, 1992; Tacarindua et al., 2013; Nakagawa et al., 2020). The impact of high temperatures on soybean yield and quality has been studied using open-enclosure facilities, controlled artificial climate chambers, or crop models based on historical data (Gibson, 1992; Tacarindua et al., 2013; Nakagawa et al., 2020). A negative correlation between soybean yield and seasonal warming has been discovered (Kucharik and Serbin, 2008). For every 1°C increase, soybean yield decreases by an average of 17% (Lobell and Asner, 2003). The high temperature associated with damage to soybean growth and development took place in controlled chambers and greenhouses (Gibson, 1992; Tacarindua et al., 2013; Nakagawa et al., 2020). Some studies reported a decrease in yield at high temperatures in the soybean seed filling stage (Gibson and Mullen, 1996; Siebers et al., 2015). It was observed that high temperature stress experienced during mid-reproductive growth is more detrimental to seed yield and seed size than that experienced during early or late reproductive development stage (Puteh et al., 2013). Extreme heat or heat stress can cause irreversible damage to soybean growth and development, and can even cause death (Feng et al., 2020). The moderately high temperatures are more likely to lead to morphological, structural, physiological, and biochemical changes in the thermal adaptation to warming conditions (Puteh et al., 2013). It was found that moderate high night temperature of 29°C reduced photosynthetic rates and pollen germination, resulting in decreased pod numbers and seed weight (Djanaguiraman et al., 2013). However, the effect of high night temperature, seed’s C and N accumulation, and corresponding underlying mechanisms in soybeans remains unclear.

It has been suggested that high night temperature stimulate carbohydrate usage (Loka and Oosterhuis, 2010). The yield decline is mainly due to a reduction in the biomass allocated to the coffers and a decrease in the content of dry matter and its synthetics. Losses caused by high night temperature are usually accompanied by a decline in biomass accumulation (Zhang et al., 2013; Jing et al., 2016). Daytime is a crucial period for photosynthesis in soybeans; at night, the plants turn into mere consumers of the bioproducts of photosynthesis, which accumulate during the day (Xu et al., 2021). Many studies have confirmed that respiration rates of many crops such as rice (Alvarado-Sanabria et al., 2017), wheat and barley (García et al., 2015), maize (Kucharik and Serbin, 2008), and cotton (Loka and Oosterhuis, 2010) increase at high night temperature. The same result was also reported for soybeans (Bunce, 2005; Djanaguiraman et al., 2013; Lin et al., 2021). As plants enhance nocturnal respiration (Rn), more photosynthetic bioproducts are consumed, thereby reducing the sugar reserves available for seed filling (Xu et al., 2021), night temperatures of 10°C above ambient lead to a 2.6-fold enhancement in rapid nocturnal respiration. However, the response diminishes after thermal acclimation and causes yield loss (Peraudeau et al., 2015). It was reported that the high night temperature resulted in the higher Rn and decline of the yield in soybean by 4.6% per °C (Lin et al., 2021). Another study found that various soybean varieties showed different level of tolerance to high temperature, only a few varieties showed decrease yield under high night temperature (Shu, 2021). One study has also observed that high night temperature causes many environmental variables and that respiration has no effect on yield or biomass (Frantz et al., 2004).

Non-structural carbohydrates in leaves, such as sucrose and starch, represent the state of photosynthesis in the leaf, provide whole-plant energy, and contribute to the accumulation of structural carbohydrates (Hussain et al., 2019; Stitt et al., 1987). Leaf senescence may affect crop yield in two different ways. On one hand, delayed leaf senescence can increase yield by prolonging photosynthesis and extending seed filling period; On the other hand, delayed leaf senescence may affect nutrient reuse to the sink organ, which may harm yield (Guo et al., 2021). Sucrose is the main source of carbon energy for developing seeds and sucrose transport is the central system of carbon resource allocation for the whole plant (Ruan, 2014; Petridis et al., 2020; Zhang et al., 2016). It has been reported that sucrose content decreases with the increase in temperature (Loka and Oosterhuis, 2010). Studies have shown proteomic changes in different tissues like leaves and anthers of plants at high temperatures, as well as differential expression of proteins related to various physiological processes, such as carbohydrate metabolism (Das et al., 2015; Kumar et al., 2022; Zhou et al., 2017). The balance between sucrose synthesis and catabolism in leaves is mediated by influencing the activity of sucrose synthase (SS) and sucrose phosphate synthase (SPS), two key enzymes that also control the biosynthesis of carbon into starch (Stitt and Zeeman, 2012) However, few researchers have studied the effect of night temperature on the biosynthesis of leaf sucrose.

Currently, RNA-seq technology is widely used in temperature regulation studies in plants (Gillman et al., 2019; Shu et al., 2020). RNA-Seq analysis of whole wheat kernels showed that high temperature induced down-regulation of genes involved in regulating pericarp cell wall expansion (Kino et al., 2020). This down-regulation coincided with a reduction in total grain water content and weight after the same treatment period, supporting the theory that high temperature may lead to a reduction in mature grain weight (Kino et al., 2020). Proteins like heat shock proteins also known to play a key role in leguminous plants like Vicia faba and other plants under heat stress (Kumar et al., 2015; Kumar et al., 2016; Kumar et al., 2020; Tiwari et al., 2021).

The aims of the study were (i) to evaluate the effect of high night temperature during the seed filling stage on the yield and seed compositions, (ii) to discover the dynamic changes of photosynthetic physiological characteristics, dry matter accumulation and NSC linked to stress tolerance triggered by high night temperature, and (iii) to analyze the molecular basis of changes in the leaves, specifically sugar metabolism.

Soybean is sensitive to light and temperature. When subjected to photothermal change, it must constantly employ adaptive mechanisms to maintain homeostasis in photosynthesis, carbon fixation, and photosynthetic production (Pommerrenig et al., 2018). These adaptive mechanisms can both ensure the normal growth of the soybean but caused changes of biomass accumulation, individual development, material partitioning, and yield (Du et al., 2020). A systematic understanding of the relationship between elevated night temperatures and yield-related physiological indicators is beneficial for improving field cropping strategies and achieving improved soybean yield and quality.

The negative effect of high night temperature on soybean yield was reported in previous studies (Egli, 2010; Nakagawa et al., 2020), and it was consistent with the results of many other crops such as rice (Bahuguna et al., 2016, Welch et al., 2010), wheat (García et al., 2015; García et al., 2016), and cotton (Loka and Oosterhuis, 2010). It was found that high night temperature increased respiration-driven carbon losses and hindered the transport of photosynthetic assimilation products from source organs to reservoir, resulting in inadequate seed filling and ultimately a reduction in yield, which is consistent with the study of Tacarindua et al. (2013). The source-sink relationship during grain development was affected by temperature was also studied in many crops, such as rice (Arshad et al., 2017) and wheat (Hein et al., 2022). The negative correlation of soybean yield and quality under high night temperature reported in previous study (Choi et al., 2016) was confirmed in our study. It has also been reported that high temperature may affect seed weight by altering protein content (Nakagawa et al., 2020). A “concentration effect” resulted from a decline in the proportion of carbohydrate and seed size, caused the increase relative protein and oil content was also observed in our study. Furthermore, the protein and oil yield of individual soybean plants was found reduced under high night temperature, implying the high night temperature affects the accumulation of different components, but it had more effects on the carbohydrates than proteins and oils.

In this study, 28°C high night temperature treatment led to an increase in net photosynthetic rate in early 7 DAT functional leaves, contrary to the results of some studies of heat stress (Jumrani et al., 2016; Prasad et al., 2017; Ferguson et al., 2021), this may as a result of treatment carried out in the night and the 28°C night temperature was a mild heat stress for soybean (Hatfield et al., 2011). Some studies investigating the effects of night temperature concluded that there was no effect on net photosynthetic rate (Zhang et al., 2020), which was also different from our results. This may be due to the differences of treatment and crop species. Previous studies have reported that soybean leaves senesce later in controlled temperature treatments with 1-3°C higher environmental temperature (Tacarindua et al., 2013). This is consistent with the findings of our study and the stay-green of the leaf could be the reason for the decrease translocation of accumulated photosynthetic assimilation products in the leaves and the weakened efficiency of the photosynthetic product utilization.

Elevated levels of sucrose occurred early in this study, which may be related to the compensation for the carbon starvation caused by its accelerated degradation at night (Peraudeau et al., 2015; MacNeill et al., 2017). After a period of treatment, the sucrose content begins to remain relatively stable as it adapts to the change in temperature. The starch content in stems and leaves was found to be lower than the control under the high night temperature treatment after 21 DAT. The decrease in sucrose content in the seeds partly explains the decrease of the residue in seeds. Similarly, in rice, it was found that one of the causes for grain stunting under heat stress during reproductive growth was the restricted carbohydrate availability, and that heat stress reduced the movement of photosynthetic assimilation products into the seeds and inhibited the accumulation of starch in the seeds (Arshad et al., 2017).

The identified DEGs for photosynthetic and carbohydrate are associated with early increased sugar accumulation in the leaves treated under high night temperature treatment. Photosynthesis could be improved by the increased carbon fixation and sugar metabolism. Also, leaf senescence triggered by sugar accumulation leads to increase of nitrogen content (Wingler et al., 2006, Kaschuk et al., 2010). DEGs related to the transported nitrogenous compounds glutamine and glutamate were identified (Masclaux-Daubresse et al., 2010). Some amino acids or organic acids, such as proline and glutamate, are synthesized in large quantities when plants are subjected to heat stress (Vergara-Diaz et al., 2020; Wahid, 2006), and we also find the DEGs for these compounds. The transcriptome results reflect the response mechanism to high night temperature treatment conditions at the beginning of the seed filling stage. Some previous studies have suggested that changes in seed size and quality under heat treatment conditions could be associated with the expression of some heat-stimulated proteins (Kumar et al., 2015; Kumar et al., 2016; Kumar et al., 2020; Tiwari et al., 2021). Most HSP-related genes showed down-regulation in our study ( Table S1 ), it could be explained by the mild stress caused by the moderate high night temperature we used.

This study expounds on the current understanding of the relationships between photosynthesis, plant development, carbon and nitrogen transport, and metabolism under high night temperature conditions. In addition to exploring response and tolerance mechanisms at the molecular level, we also need to optimize crop management strategies to improve soybean plants’ resistance to nighttime warming. In the context of global warming, breeding soybean varieties with low respiration and high energy efficiency is one of the most important ways to effectively address the world food crisis.

This study indicated that high night temperature during the soybean seed filling period (R5-R7) reduced seed yield and changed the proportion of the seed compositions. The decline of carbohydrates in seeds was the primary reason for the reduction in yield. The dynamic changes of dry matter accumulations, non-structural carbohydrates, net photosynthetic rate and SPAD value, as well as the transcriptome analysis explained the adaptive mechanisms for soybean to high night temperature stress at physiological and molecular level. The study is of significance for improving soybean breeding and cultivation practice to fight the global warming.

The original contributions presented in the study are publicly available. This data can be found here: NCBI, PRJNA892245 and SAMN31361599-31361604.

LY and WS conceived the initial research plan, conducted the experiments, and analyzed the data. CW supervised the research and provided the laboratory infrastructure and funding. LY prepared the graphs and wrote the article. CW, DJ, WS, CX, and ES critically reviewed and edited the manuscript. All authors contributed to the article and approved the submitted version.