Wenhui Zheng ·Renshan Li·Qingpeng Yang ·Weidong Zhang ·Ke Huang·Xin Guan·Longchi Chen·Xin Yu·Qingkui Wang·Silong Wang
Abstract Stored nonstructural carbohydrates (NSC) indicate a balance between photosynthetic carbon (C) assimilation and growth investment or loss through respiration and root exudation.They play an important role in plant function and whole-plant level C cycling.CO2 elevation and nitrogen (N) deposition,which are two major environmental issues worldwide,affect plant photosynthetic C assimilation and C release in forest ecosystems.However,information regarding the effect of CO2 elevation and N deposition on NSC storage in different organs remains limited,especially regarding the trade-offbetween growth and NSC reserves.Therefore,here we analyzed the variations in the NSC storage in different organs of Chinese fir (Cunninghamia lanceolata) under CO2 elevation and N addition and found that NSC concentrations and contents in all organs of Chinese fir saplings increased remarkably under CO2 elevation.However,N addition induced differential accumulation of NSC among various organs.Specifically,N addition decreased the NSC concentrations of needles,branches,stems,and fine roots,but increased the NSC contents of branches and coarse roots.The increase in the NSC contents of roots was more pronounced than that in the NSC content of aboveground organs under CO2 elevation.The role of N addition in the increase in the structural biomass of aboveground organs was greater than that in the increase in the structural biomass of roots.This result indicated that a different tradeoffbetween growth and NSC storage occurred to alleviate resource limitations under CO2 elevation and N addition and highlights the importance of separating biomass into structural biomass and NSC reserves when investigating the effects of environmental change on biomass allocation.
Keywords Biomass partition·CO2 elevation·N deposition·Nonstructural carbohydrates·Structural biomass
A fluctuation of nonstructural carbohydrates (NSC) in plant organs represents an asynchrony between carbon (C) supply and C demand (Kobe et al.2010;Sala et al.2012;Hartmann and Trumbore 2016).NSC are primarily derived from foliage photosynthates (Kozlowski 1992;Carbone et al.2013) and translocate to other organs for energy metabolism,osmoregulation,and defense compound synthesis (Hartmann and Trumbore 2016).Given that NSC constitute 8%-17% of a tree’s biomass (Wurth et al.2005;Mei et al.2015;Martínez-Vilalta et al.2016),they are a vital component in terrestrial C cycling.Furthermore,NSC dynamics across plant organs ref lect C source-sink status and organ function in adapting to environmental changes (Hartmann and Trumbore 2016).Despite its vital role in plant function and in stand-level terrestrial C cycling,NSC allocation of plant organs to environment stresses has been the focus of very limited research.With the current and expected changes to the global climate,however,a comprehensive understanding of NSC allocation in response to environmental variations is essential (Li et al.2013;McDowell et al.2008;Yang et al.2016).
Because atmospheric CO2concentration has increased fro m 280 μmol mol-1before the Industrial Revolution to 414 μmol mol-1in 2020 (Dong et al.2021),the impacts of CO2elevation on C cycling of forest ecosystems have received considerable attention (Oren et al.2001;Hamilton et al.2002;Norby et al.2010).For instance,in a sweetgum forest,Norby et al.(2002) found that CO2elevation leads to differential increases in the biomass of various organs,but NSC allocation to different organs in response to CO2elevation has been rarely investigated (Li et al.2018b).When CO2is elevated,C assimilation increases as conf irmed by an upregulation of leaf photosynthetic capacity (Arp and Drake 1991;Long et al.2004) .Thus,under CO2elevation,it is reasonable to infer that much more NSC are stored in leaves.The translocation of carbohydrates to other organs to support organ growth also increases as the photosynthetic rate increases (Li et al.2018a).Nevertheless,additional tree growth is likely limited by nutrient supply,particularly nitrogen or phosphorus,when CO2is elevated (Oren et al.2001;Norby et al.2010).In response to nutrient limitation,plants may increase fine root production and mycorrhizal infection to intensify soil exploration (Zak et al.2000;Luo et al.2004).To support root and mycorrhizal growth,additional photosynthetic C might be partitioned to underground organs.Previous studies have mostly focused on NSC concentration responses to CO2elevation at the leaf or whole-plant level (Smart et al.1994;Cheng et al.2004;Mašková et al.2017).Therefore,whether the NSC contents and concentrations in various organs vary in response to CO2elevation should be clarif ied.
Nitrogen (N) deposition is another serious global problem that is intensifying in China.N deposition rate in China increased by 0.41 kg ha-1a-1between 1980 and 2010,reaching 21.1 kg ha-1a-1in 2010 (Liu et al.2013).N deposition leads to an increase in NSC accumulation (Liu et al.2016) by promoting photosynthetic C assimilation (Evans and Terashima 1988;Nakaji et al.2001;Cechin and Fumis 2004;Granath et al.2009).It may also intensify NSC consumption through several mechanisms.First,N deposition may amplify N assimilation and consequently elevate NSC consumption by increasing the amount of energy and C skeletons consumed in N assimilation (Invers et al.2004;Monson et al.2006).Second,during the stimulation of plant growth after N deposition,more carbohydrates may be allocated to structural biomass (Liu et al.2 016).Although previous studies have reported that N assimilation rates of leaves and roots show different responses to N addition (Invers et al.2004),the changes in NSC consumption and allocation in various organs under N addition remain unclear.Thus,clarifying NSC dynamics in different organs under increased N deposition is urgently needed.
The allocation of biomass,which comprises structural biomass and NSC reserves,is a vital strategy used by plants to adapt to environmental conditions (Guo et al.2016).Separating biomass into structural biomass and NSC storage is critical because the allocation of structural biomass and NSC reserves is the result of the trade-offbetween growth and carbohydrate storage (Canham et al.1999; Kobe et al.2010).Although the allocation of biomass under elevated CO2and N has received attention (Hättenschwiler and Körner 1997;Norby et al.2002),few works have investigated the allocation of the separate biomass components at the whole-plant level under CO2elevation and N deposition.In addition,whether structural biomass or NSC reserves is the major contributor to alleviate resource limitations remains unclear.Optimal partitioning theory states that plants can increase biomass allocation to organs to acquire the resource(s) that most limits their growth (Bloom et al.1985).Kobe et al.(2010),however,reported that incremental increases in NSC content rather than in structural biomass,are mainly responsible for the increase in biomass allocation to roots under nutrient limitation.Therefore,the responses of allocation of structural biomass and NSC content to CO2elevation and N deposition require clarification.
Chinese fri [Cunninghamia lanceolata(Lamb.) Hook.] is widely planted in southern China and plays a vital role in timber production and C sequestration (Wang et al.2 017).Thus,further insight into the responses of NSC allocation in Chinese fir to CO2elevation and N deposition are needed to design strategies to address the effects of global climate change.Here we (1) investigated the responses of NSC concentrations and contents of various organs to CO2elevation or N deposition,and (2) compared the resultant changes in structural biomass and NSC storage among aboveground organs and roots.
Study site and experimental design
The experiment was conducted at Huitong National Research Station of Forest Ecosystem (26°50＇ N,109°36＇ E) in Hunan Province,China.This region has a typical humid mid-subtropical monsoon climate that is characterized by a warm and humid summer and a relatively dry and cold winter.The annual average air temperature is 16.5 °C,and the monthly mean maximum and minimum air temperatures of 29.0 °C and 1.9 °C occur in July and January,respectively.Annual precipitation ranges from 1200 to 1400 mm,mainly during April to July.
In February 2016,50 one-year-old Chinese fir saplings were planted in plastic pots (25 cm diameter,50 cm height) filled with surface soil (0-30 cm) that had been homogenized after its removal from a Chinese fri plantation.All pots were periodically watered to 60% field capacity to replace water lost via evapotranspiration.
In August 2016,36 Chinese fir saplings with similar basal diameters and heights were selected and separated randomly into six equal groups.Each group was transferred to a randomly assigned closed climate-controlled chambers.From 06:00 to 18:00 every day,three chambers received ambient air CO2concentration (approximately 400 μmol mol-1) and the other three elevated CO2concentration (600 ± 50 μmol mol-1) that was controlled using a CO2detection system (Shsen-QZD,Institute of Shengsen Numerical Control Technology,Qingdao,China).Three Chinese fir saplings in each chamber were randomly chosen as parallel samples to receive N fertilizer in the form of NH4N O3at the rate of 10 g N m-2a-1via irrigation once a month.The three other saplings received an equal amount of deionized water.Soil moisture was maintained at approximately 60% of field capacity during the experiment.Thus,four treatments were imposed:(1) ambient air CO2concentration and no N (EC0N0),(2) ambient air CO2concentration and N addition (EC0N1),(3) elevated CO2concentration and no N (EC1N0),and (4) elevated CO2concentration and N addition (EC1N1).
Sample collection and NSC measurement
In September 2017,all saplings in the chambers were harvested and separated into different organ types (needles,branches,stems,and roots).Needles and branches were each separated into current and previous year groups.Roots were separated into coarse (diameter ≥ 2 mm) and fine (diameter ＜ 2 mm) roots.To reduce variability,we prepared mixed samples by combining the parallel samples from each chamber on the basis of organ types.All samples from different organs were oven-dried at 80 °C to a constant mass then weight to estimate biomass.The samples were then milled to a fine powder to measure NSC using the method described by Li et al.(2018a) and Yang et al.(2016):Soluble sugars (i.e.,glucose,sucrose,and fructose) and starch concentration were measured and added to estimate total NSC concentration.
The NSC content for each organ was calculated as Organ-specific concentration × Organ-specific biomass.Structural biomass was then calculated as Biomass minus NSC content similar to the methods of Canham et al.(1999) and Kobe et al.(2010).
Statistical analyses
The effects of CO2elevation,N addition,and their interaction on concentrations and contents of NSC,concentrations and contents of starch and of soluble sugars in stems,coarse roots,and fine roots were all evaluated using a two-way ANOVA implemented in R version 4.1.3 (R Core Team 2022).The mixed effects model was applied to evaluate the effect of CO2elevation,N addition,needle age or branch age,and their interactions on NSC concentrations and NSC contents in needles and branches.This model was run using the lmer function in R by loading the lme 4 and lmer Test packages.The ANOVA function in R was used for parametric tests.In this model,CO2elevation,N addition,and needle age or branch age were used as fixed effects;the chamber was used as a random effect.The significance level was set atP＜ 0.05 for all analyses unless otherwise stated.
Concentrations of NSC and its two fractions in different organs
Needle NSC (Fig.1 a,P=0.006),soluble sugars (Fig.1 c,P=0.028),and starch (Fig.1 e,P=0.002) concentrations greatly decreased with needle age.CO2elevation remarkably increased needle NSC concentrations (Fig.1 a,P=0.001) and its two fractions (soluble sugars (Fig.1 c,P ＜ 0.001) and starch (Fig.1 e,P=0.029).However,a marginally signif ciant decrease in needle NSC concentration (Fig.1 a,P=0.057) and a significant decrease in needle starch concentration (Fig.1 e,P ＜ 0.001) were found under N addition.The soluble sugars concentration in needles did not change in response to N addition (Fig.1 c,P=0.804).N addition and needle age exerted an interactive effect on starch concentration (Fig.1 e,P=0.022).This effect indicates that the decrease in the starch concentration of current year needles after N addition was greater than the decrease in the previous year needles (Fig.1 e).
Fig.1 NSC (a,b),soluble sugars (c,d),and starch (e,f) concentrations in needles (a,c,e) and branches (b,d,f) from Chinese fir saplings under CO2 elevation and N deposition.EC0:ambient air CO2 concentration;EC1:elevated CO2 concentration;N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Different lowercase letters denote significant differences between EC1N0 and EC1N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05,** P ＜ 0.01.Values are expressed as means ± SE (n =3)
Unlike the NSC concentration in needles,the concentration of NSC and its two fractions in branches did not differ based on branch age (Fig.1 b,d,f,P＞ 0.05).CO2elevation significantly increased branch NSC (Fig.1 b,P＜ 0.001),soluble sugars (Fig.1 d,P＜ 0.001),and starch (Fig.1 f,P＜ 0.001) concentrations.Conversely,N addition significantly decreased branch NSC (Fig.1 b,P=0.001),soluble sugars (Fig.1 d,P=0.007),and starch (Fig.1 f,P=0.005) concentrations.CO2elevation and branch age had an interactive effect on branch NSC (Fig.1 b,P=0.004) and soluble sugars concentrations (Fig.1 d,P=0.002).These results demonstrate that the increase in NSC and soluble sugars concentrations in current year branches was more prominent than in previous year branches under CO2elevation.
CO2elevation significantly increased the concentrations of NSC (Fig.2 a,P ＜ 0.001),soluble sugars (Fig.2 b,P ＜ 0.001),and starch (Fig.2 c,P=0.004) in stems,whereas N addition signif ciantly decreased the concentrations of NSC (Fig.2 a,P=0.009) and soluble sugars (Fig.2 b,P=0.018).CO2elevation and N addition exerted a significant interactive effect on NSC (Fig.2 a,P=0.025) and soluble sugars concentrations (Fig.2 b,P=0.005) in stems.This effect showed that the decrease in stem NSC and soluble sugars concentrations with N addition only happened in ambient CO2,not in elevated CO2.
Fig.2 NSC (a),soluble sugars (b),and starch (c) concentration in stems from Chinese fir saplings under CO2 elevation and N deposition.EC0:ambient air CO2 concentration;EC1:elevated CO2 concentration;N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05,** P ＜ 0.01.Values are expressed as means ± SE (n =3)
CO2elevation significantly increased the starch (Fig.3 e,P＜ 0.001) and NSC concentrations (Fig.3 a,P＜ 0.001) in coarse roots and starch (Fig.3 f,P＜ 0.001) and NSC (Fig.3 b,P＜ 0.001) concentrations in fine roots.However,the NSC,soluble sugars,and starch concentrations in coarse roots did not change in response to N addition (Figs.3 a,c,and e,P＞ 0.05).N addition significantly decreased the NSC (Fig.3 b,P=0.004) and soluble sugars concentrations (Fig.3 d,P=0.001) in fine roots.A significant interactive effect between CO2elevation and N addition on the NSC concentration of coarse roots was found (Fig.3 a,P=0.041).
Contents of NSC and its two fractions in different organs
Similar to needle NSC concentration,the contents of needle NSC and its two fractions decreased with needle age (Figs.4 a,c,e,P ＜ 0.001).Similarly,branch NSC (Fig.4 b,P=0.045) and soluble sugars (Fig.4 d,P=0.047) contents decreased with branch age.With the exception of soluble sugars content in fine root,NSC,soluble sugars and starch contents significantly increased in all organs under CO2elevation (Figs.4,5,and 6,P＜ 0.05).However,NSC content and its two fractions in various organs varied in their responses to N addition.In needles,N addition significantly increased soluble sugars content (Fig.4 c,P=0.005) but significantly decreased starch content (Fig.4 e,P=0.040).In branches,NSC (Fig.4 b,P=0.007) and soluble sugars (Fig.4 d,P=0.010) contents significantly increased with N addition,as did starch content (Fig.6 e,P=0.003) and NSC content (Fig.6 a,P=0.003) in coarse roots.However,soluble sugar content in fine roots significantly decreased with N addition (Fig.6 d,P=0.036).CO2elevation and N addition had significant interactive effects on needle NSC (Fig.4 a,P=0.004) and soluble sugars (Fig.4 c,P=0.002),branch NSC (Fig.4 b,P=0.003) and soluble sugars (Fig.4 d,P=0.006),stem NSC content (Fig.5 a,P=0.049),and on coarse-root starch (Fig.6 e,P=0.003) and NSC content (Fig.6 a,P=0.003).CO2elevation and needle age had a significant interactive effect on needle NSC (Fig.4 a,P＜ 0.001),starch (Fig.4 e,P＜ 0.001) and soluble sugars content (Fig.4 c,P＜ 0.001).
Fig.3 NSC (a,b),soluble sugars (c,d),and starch (e,f) concentrations in coarse (a,c,e) and fine roots (b,d,f) from Chinese fir saplings under CO2 elevation and N addition.EC0:ambient air CO2; EC1:elevated CO2; N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05,** P ＜ 0.01.Values are expressed as means ± SE (n =3)
Fig.4 NSC (a,b),soluble sugars (c,d),and starch (e,f) contents in needles (a,c,e) and branches (b,d,f) from Chinese fir saplings under CO2 elevation and N addition.EC0:ambient air CO2 concentration;EC1:elevated CO2 concentration;N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Different lowercase letters denote significant differences between EC1N0 and EC1N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05,** P ＜ 0.01.Values are expressed as means ± SE (n =3)
Fig.5 NSC (a),soluble sugars (b),and starch (c) contents in stems from Chinese fir saplings under CO2 elevation and N addition.EC0:ambient air CO2 concentration;EC1:elevated CO2 concentration;N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05,** P ＜ 0.01.Values are expressed as means ± SE (n =3)
Fig.6 NSC (a,b),soluble sugars (c,d),and starch (e,f) contents in coarse (a,c,e) and fine roots (b,d,f) from Chinese fir saplings under CO2 elevation and N addition.EC0:ambient air CO2 concentration;EC1:elevated CO2 concentration;N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Different lowercase letters denote significant differences between EC1N0 and EC1N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05,** P ＜ 0.01.Values are expressed as means ± SE (n =3)
Comparison of allocation of structural biomass and NSC content to aboveground organs and roots
Under CO2elevation,NSC content was preferentially allocated to roots over aboveground organs (Fig.7 a,P=0.027),but the proportion of the structural biomass of aboveground organs to the structural biomass of roots did not change in response to CO2elevation (Fig.7 b,P=0.139).By contrast,N addition promoted the preferential partitioning of structural biomass to aboveground organs instead of underground roots,as indicated by the significant increment in the structural biomass of aboveground organs relative to that of roots (Fig.7 b,P=0.008).The proportion of the NSC content of aboveground organs to the NSC content of roots did not change in response to N addition (Fig.7 a,P=0.473).
Fig.7 Ratio of the sum of NSC content of aboveground (AG) organs to the NSC content in roots (a) and ratio of the sum of structural (struct) C content of aboveground organs to the structural C content in roots (b) from Chinese fri saplings under CO2 elevation and N deposition.EC0:ambient air CO2 concentration;EC1:elevated CO2 concentration;N0:no N;and N1:N addition.Different uppercase letters denote significant differences between EC0N0 and EC0N1 (P ＜ 0.05).Different lowercase letters denote signif ciant differences between EC1N0 and EC1N1 (P ＜ 0.05).Significant differences between EC0N0 and EC1N0 or between EC0N1 and EC1N1 are denoted by asterisks:* P ＜ 0.05.Values are expressed as means ± SE (n =3)
Effect of CO2 elevation on NSC concentration and content of different organs
The positive effect of CO2elevation on NSC accumulation in needles could be due to the following reasons.First,NSC are mainly produced in leaves through photosynthetic C assimilation (Kozlowski 1992),the upregulation of leaf photosynthetic capacity under CO2elevation increases C assimilation (Arp and Drake 1991;Long et al.2004;Luo et al.2006).Second,CO2elevation is likely to reduce needle respiration rates (Idso and Kimball 1992;Zha et al.2002;Long et al.2004).Thus,the increase in C sources and the reduction in C sinks resulted in needle NSC accumulation under CO2elevation.In addition,the response of NSC accumulation to CO2elevation was dependent on needle age;the accumulation in current year needles was more pronounced than in previous year needles (Fig.4 a,c,e) because photosynthetic rate in current year needles is more sensitive than in previous year needles to CO2elevation (Tissue et al.2001).
CO2elevation increased branch and stem NSC concentration and content by augmenting the translocation of photosynthates from needles to branches and stems.We found that CO2elevation promoted sapling growth (Fig.S2) as reported previously (Curtis and Wang 1998;De Souza et al.2008).Therefore,the excess photosynthate fixed by needles is translocated to branches and stems to promote their elongation and maximize space occupation and light harvest (Li et al.2018a).Second,the additional growth of biomass under CO2elevation increased the amounts of soil nutrients required for organic matter production (Huang et al.2007).Thus,large amounts of NSC must be translocated to branches and stems for nutrient transport given their vital roles in phloem transport (Dietze et al.2014).
Similar to the accumulation in aboveground organs,NSC accumulation in coarse and fine roots increased under CO2elevation.This response may be related to the nutrient limitations experienced by plants under CO2elevation (Fig.S3).Plants could alleviate nutrient limitation by amplifying their exploration of soil for available nutrients (Luo et al.2004).Thus,the NSC fixed by needles were translocated to underground organs to support the root activity and growth needed for nutrient acquisition (Kobe et al.2010).Interestingly,starch,rather than soluble sugars,was mainly responsible for the increase in NSC accumulation in roots under CO2elevation.This result may be attributed to the different functions between starch and soluble sugars.Starch is used as a carbohydrate reserve and represents a relatively recalcitrant C pool (Hartmann and Trumbore 2016).Meanwhile,soluble sugars are usually used for immediate functions (e.g.,substrates for respiration,osmoregulation and transport compounds) and represent a short-term pool (Du et al.2020).Therefore,part of the photosynthate allocated to roots was converted to starch for future use,resulting in starch accumulation.Plants exchange soluble sugars,especially glucose,for nutrients with symbionts under CO2elevation;thus,soluble sugars are maintained at a critical level (Nehls et al.2010;Smith and Smith 2011;Martínez-Vilalta et al.2016).
Interestingly,CO2elevation resulted in the preferential allocation of NSC content to roots instead of to aboveground organs.The allocation of structural biomass to aboveground organs vs roots,however,did not change in response to CO2elevation.The promotion of plant growth under CO2elevation may result in nutrient limitation,especially nitrogen and phosphorus (Oren et al.2001;Norby et al.2010),because nitrogen and phosphorus concentration in Chinese fir saplings decreased when CO2was elevated (Fig.S3).According to the optimal partitioning theory,biomass is preferentially allocated to roots to increase the absorptive surface area of roots as the main mechanism for plants to capture limited sources (Bloom et al.1985;Zak et al.2000;Luo et al.2004).However,the present study found that only the NSC reserves of roots increased under nutrient limitation.Similarly,Kobe et al.(2010) found that enhanced NSC storage contributed more than increased structural biomass to the change in root biomass under nutrient limitation.This result is probably related to the fact that,in our study high NSC reserves provide more advantages than just an increase in absorptive surface area to gain more nutrients.First,root NSC provide energy to roots and mycorrhizae for nutrient uptake and are a carbon source in rhizosphere exudates that prime microbial populations to mineralize organic matter and obtain nutrients (Rothstein et al.2000;Dijkstra and Cheng 2007).Second,NSC storage,not the immediate construction of fine roots,favors plant survival when nutrients are scarce because newly formed fine roots consume energy required for respiration (Kobe et al.2010).
Effect of N addition on the NSC concentration and content of different organs
Various factors could contribute to the decreased starch and NSC concentrations in needles under N addition (Fig.1).First,the allocation of NSC for N assimilation needed to synthesize proteins likely increased under N addition.Kraemer et al.(1997) reported that high N assimilation rates is associated with high N contents.N addition increased foliar N concentration in our study (Fig.S3) and in others (Hicks et al.2000;Mo et al.2008;Lü et al.2016).Therefore,NSC consumption goes up due to the increased demand for C skeletons and energy for N assimilation (Foyer et al.2003;Cheng et al.2004;Invers et al.2004).Second,because N addition promotes plant growth (Cechin and Fumis 2004;Mo et al.2008),photosynthate allocation to structural biomass increased after N addition,resulting in the decrease in needle NSC concentration (Liu et al.2016).This interpretation is well supported by the increase in sapling structural biomass under N addition in our study (Fig.S4).
Branch and stem NSC concentrations decreased under N addition,whereas NSC content increased or remained stable.Similar to the decrease in needle NSC concentration,the decrease in branch and stem NSC concentrations may be attributed to increased use of NSC for N assimilation,as indicated by the increase in the total N concentrations of branches and stems under N addition (Fig.S3).Moreover,the promotion of branch and stem growth after N addition was responsible for the reduction in NSC concentration.This interpretation is in line with the increases in branch (Fig.S1) and stem (Fig.S2) biomass under N addition.The NSC contents in different organs were calculated as organ-specif ci concentrations multiplied by organ-specific biomass.Therefore,the decrease in NSC concentration under N addition may be used to promote growth of branch and stem and result in increased or constant NSC contents.
N addition decreased the NSC concentration of fnie roots,but the NSC concentration of coarse roots was not responsive to N addition.The different physiological functions of coarse and fine roots may account for the different responses in NSC concentrations after N addition.Coarse roots play an important role in resource reserves and transport (Guo et al.2004;Hartmann and Trumbore 2 016).Thus,stable NSC reserves in coarse roots favor the survival of plants during environmental change.Fine roots are mainly responsible for nutrient uptake (Mei et al.2 015).Based on the optimal partitioning theory,N addition alleviates nutrient limitation,so that plants do not have to increase fine root production and promote mycorrhizal infection (Zak et al.2000;Luo et al.2004).Thus,the allocation of NSC to fine roots is reduced (Kobe et al.2010).Another explanation is that the N content in fine roots is higher than in coarse roots (Fig.S3) (Guo et al.2004).Root respiration rate is strongly and positively correlated with N content (Burton et al.2012;Mei et al.2015).Thus,compared with coarse roots,the much higher level of NSC in fine roots is likely used to maintain respiration.
The proportion of the NSC content in aboveground organs to the NSC content of roots was not responsive to N addition.However,we found that N addition increased structural biomass more in aboveground organs than in root,possibly because the increase in aboveground structural biomass rather than an increase in NSC reserve was conducive to alleviating the C limitation caused by N addition.Previous studies have demonstrated that the stimulation of plant growth and N assimilation by N addition could result in a C limitation (Invers et al.2004;Monson et al.2006;Liu et al.2016).Bloom et al.(1985) reported that plants may maximize their net photosynthetic rates through the timely reinvestment of carbohydrates into the continuous production of new leaves,so that the plant can enhance photosynthate acquisition.Similarly,in our study,photosynthates were mainly used for needle production,branch elongation,and stem growth to optimize light harvest by maximizing spatial occupation rather than being stored as NSC (Imaji and Seiwa 2010).
In our study of organ-specific NSC responses to CO2elevation and N addition,CO2elevation promoted NSC accumulation in all organs,and NSC contents were preferentially allocated to roots rather than to aboveground organs.This allocation pattern may be related to nutrient limitation experienced by plant under CO2elevation and the increased use of NSC as an energy source by roots to increase nutrient absorption.N addition reduced NSC concentrations in needles,branches,stems,and fine roots and increased the NSC contents in branches and coarse roots.On a wholetree basis,structural biomass was preferentially partitioned to aboveground organs instead of underground roots under N addition.This partitioning pattern indicates that timely reinvestment of carbohydrate in growth rather than in NSC reserves helps alleviate C limitation under N addition.Our results indicated that different trade-offs between structural growth and NSC storage help ameliorate resource limitations under CO2elevation and N addition in Chinese fir.Our results thus show that it is important to separate biomass into structural biomass and NSC storage when analyzing the effect of environmental change on biomass allocation.
AcknowledgementsWe thank Youzhi Yu for his invaluable assistance with laboratory and field experiments.