An Environment Transitional Zone Buffers Peatlands Carbon Loss

An Environment Transitional Zone Buffers Peatlands Carbon Loss | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article An Environment Transitional Zone Buffers Peatlands Carbon Loss Liangfeng Liu, Huai Chen, Jianqing Tian, Hongjun Wang, Dan Xue, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-849966/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract An environment transitional zone (ETzone) is usually deemed as a hotspot in biogeochemical cycle, but little is known about its response to climate change. A typical ETzone develops at the subsurface of peatland after experiencing long-term water table fluctuation, characterized by alternative aerobic and anaerobic conditions. By an extensive incubation, we found that the CO 2 emission at this ETzone was 28.31 ± 3.55 μg g -1 d -1 , 41.6% and 34.4% lower than the upper (aerobic) and lower (anaerobic) layers, respectively. Moreover, with a lowest Q 10 of 1.37, its CO 2 emission was also the least warming-responsive, which could reduce 33.7% CO 2 loss in warming scenario. This result clearly revealed that the ETzone worked as a buffer to retard carbon loss, rather than a hotspot. Surprisingly, this buffer capacity of ETzone was easily collapse if being primed by fresh carbon. Therefore, maintaining a relatively stable ETzone is critical for protecting peatland carbon stock, and the priority is to block priming effect through maintaining an intact vegetation composition. Planetary Science Environmental Chemistry Environmental Policy environment transitional zone buffers peatland carbon stock biogeochemical cycle climate change intact vegetation composition Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Introduction An environment transitional zone (ETzone), which connects zones with totally different environment and biogeochemical features, exists in the interfaces along an environmental gradient, e.g., the littoral zone between aquatic and terrestrial 1 , or the active freeze-thaw layer in discontinuous permafrost 2 , or the redox interface along the soil profile of peatlands 3 . Being in the interface of two adjacent zones, the ETzone possesses the characteristics of both and is often regarded as a hotspot of biogeochemical processes 1 , e.g., the littoral zone is hotspots of CH 4 1 and CO 2 4 emission; a temperate forest transitional slopes is hotspots of CO 2 emission and CH 4 uptake 5 . In addition, owing to frequent disturbances, ETzones are also regarded as the most vulnerable part of ecosystems 6 , especially for those fragile ecosystems, such as peatlands. In peatlands, an ETzone develops during water table fluctuation 7 (Supplementary Fig. 1), which is at the subsurface of soil profile and characterized by alternatively aerobic and anaerobic environment. Above the ETzone is aerobic surface soil under permanent aerobic conditions and abundant in living plants and roots (i.e., acrotelm); below the ETzone is anaerobic deep soil perpetually water logged and entirely without fresh plant roots (i.e., catotelm) 8,9 . Adjacent to the two soil layers, the ETzone acquires most severe disturbance due to the long term fluctuation of water table, resulting in the soil biogeochemical characteristics quite different from adjacent soil 3,7 (Table 1). However, whether this ETzone is a hotspot in soil carbon loss remains uncertain, especially under the situation of intensified climate warming and priming 10-12 . Intensified climate warming is expected to increase soil microbial decomposition and carbon export, influencing peatlands soil carbon dynamics 13,14 . A non-disturbing in situ experiment showed that 1°C warming could accelerate ecosystem respiration by 52%-60%, in which at least 69% increase originated from the soil at 25-50 cm 15 . Other in situ whole soil profile warming experiments verified that subsoil is vulnerable to warming 16,17 . A lab simulating experiment showed that subsoil (> 20 cm) was responsible for 65%-70% increase of whole profile soil carbon emission caused by warming 18 . We further explored the response of soil carbon to warming of global peatlands from ETzone and the other two zones through compiling the data of CO 2 emission rate of global peatlands soil conducting by incubation under differed temperatures and ranging a depth of 0-10 cm to 100-120 cm (Supplementary Fig. 2). The three zones were defined according to the water table data of global peatlands which were also collected from vast publications (Supplementary Fig. 3). Despite the reactive characteristic of subsoil in in situ and lab experiments, we failed to observe the anticipated higher warming response and Q 10 for ETzone in the compiled data (Fig 1). Thus, by treating the peat profile as a whole and not considering the peculiarity of the ETzone at the subsurface layer 13,19 , these experiments might have reached to inconclusive results, though established an active role of subsoil of peatlands in projecting soil carbon feedbacks to climate warming 10,20 . More research is needed to understand the characteristics of the ETzone of peatlands and its response to climate warming in terms of soil carbon stability. Priming is a composite effect of enhanced soil microbial growth and activity by increased carbon availability from vegetation roots/litters, altered pH values, soil aggregation and microbial community composition 21-23 . Researches showed that soil organic carbon (SOC) respiration can be 380% to 1200% higher with the input of plant originating compounds 24 , and that such priming effect could persist over long time 25 . In peatlands, the soil is found more vulnerable to the priming effect 26,27 and the priming effect on SOC was even strengthened under intensified climate change and human activities (e.g., water table drawdown, over-grazing) 28,29 . Similar with the threat from warming, there is so far no data for the effect of priming on SOC stability at ETzone, despite the priming effect on soil carbon stability has been widely concerned 22,30,31 . In this study, in order to understand the carbon dynamics of ETzone under intensified warming and priming, the soil CO 2 emissions and the determinants of ETzone were quantified under simulating conditions on Zoige plateau, which is a typical high-altitude minerotrophic peatlands distributed on the eastern edge of the Qinghai-Tibetan Plateau and with the global importance in predicting climate. The ETzone in our peatland was classified according to the long-term water table fluctuation (Supplementary Fig. 1). Soil samples from the ETzone (subsurface: 26-75 cm) and the two adjacent layers (aerobic surface layer: 0-25 cm; anaerobic deep layer: 76-100 cm) were incubated under the pristine field conditions (surface, aerobic; subsurface, anaerobic; deep, anaerobic) with three treatments: control (mean temperature in growing season: 8°C), warming (18°C) and substrate addition (8°C). Three substrates (oxalic acid, glucose, and cinnamic acid) found primarily in plant litter/roots exudate 32 were added into soils to simulate the priming effect. The same treatments were also applied to soil samples that were incubated under aerobic conditions to simulate the future degraded scenario when the water table in peatlands decreases severely and the peat soil of both ETzone and the deep layer are exposed in aerobic environment. The CO 2 emission rate was monitored weekly to quantify the response of soil carbon to warming. The δ 13 C of respired CO 2 was analyzed before and after substrate addition to quantify the priming effect of three substrates on soil carbon stability 33 . Soil variables of carbon component (Fe bound organic carbon, Fe-OC; water-soluble phenolics: H 2 O-PhO), nitrogen nutrient (NH+ 4), microbial indexes (microbial biomass carbon: MBC; relative abundance), and enzyme activities (phenolic oxidase, peroxidase, β-1-4-glucosidase, and cellobiohydrolase) representing carbon quality, soil nutrient, and decomposers were determined during incubation to explore the main determinants of soil CO 2 emission of three layers, by piecewise structure equation model (SEM). Methods Sampling . Soil samples were collected from Ruokeba peatland on the Zoige Plateau, northeastern Qinghai-Tibet Plateau. Zoige Plateau is characterized with low temperature (mean annual temperature of -3.3°C) and rich river system that accelerates peat accumulation. Peatlands on Zoige Plateau covers an area of 4,605 km 2 , with an average peat depth of 0.2-6.0 m. The major vegetation comprises of Equisetum ramosissimun, Scirpus triqueter, and Blysmus sinocompressus. Influenced by human disturbance and climate change, water table at Ruokeba peatland decreased severely and influenced soil carbon dynamic for years. During long-term water table fluctuation, a distinct environment transitional zone (ETzone) at subsurface soil (characterized with periodically aerobic and anaerobic due to water table fluctuation) was shaped out. Soil adjacent to ETzone includes upper aerobic surface soil (characterized with predominantly aerobic, abundant in living plants and roots) and lower anaerobic deep soil (characterized with persistent water logging and lacks of plant roots). Soil cores (105 cm in depth) were sampled at three randomly selected sites in May 2015, using a vibracore with a diameter of 8 cm. All soil cores were divided into three layers according to the water table fluctuation (Supplementary Fig. 1): aerobic surface soil (0-25 cm), subsurface soil (ETzone: 26-75 cm) and anaerobic deep soil (76-100 cm). Each layer soil from the same site was mixed, sieved (2 mm) and transported to laboratory using sterile bags in ice boxes. In laboratory, soil sample was stored at 4°C for incubation and soil characteristics analysis, and at -20°C for microbial analysis. Incubation . Soil was incubated in the conditions of field (surface, aerobic; subsurface, anaerobic; deep, anaerobic), future degraded scenario (aerobic), with treatments of control, warming and substrate addition. Sieved soil (80 g) of each layer was placed in a 500-ml glass jar and sealed using a rubber stopper with two openings for gas sampling and headspace exchange. Jars headspace was replaced with N 2 or CO 2 -free air to create anaerobic and aerobic conditions, respectively 18 . In warming experiment, jars were placed at 8°C (mean temperature in growing season) as control and 18°C as warming (3 depths * 2 conditions * 2 temperatures * 3 replicates = 36 jars). After pre-incubation for seven days, the headspace gas was weekly sampled to measure CO 2 emission rate. At 12 hours before each sampling, the headspace was flushed with N 2 or CO 2 -free air again to remove the accumulated CO 2 . The headspace gas was sampled by vacuum tube and CO 2 concentration was measured by gas chromatography (Agilent 7890A, Agilent Co., USA) with a flame ionization detector (FID) operating at 250°C. Soil water content was kept stable during incubation by constant weight and all jars were incubated for 154 days to evaluate long-term CO 2 emission. In substrate addition experiment, two sets of prepared jars as in warming experiment were incubated in aerobic and anaerobic conditions for 154 days to exhaust most labile carbon. Then three substrates with special 13 C were added into soil in powder: oxalate with δ 13 C of -0.78 (AR, ≥ 99.5%, Macklin), glucose with δ 13 C of -12.43 (labeled glucose (U- 13 C, 99%, CIL) and glucose (AR ≥ 99.5%, Macklin) mixed by weight ratio of 1:1 ), and cinnamic acid with δ 13 C of -5.25 (AR, ≥ 99.5%, Macklin). The δ 13 C of each substrate was determined before being added into soil (3 substrates * 3 depth * 2 conditions *3 replications = 54 jars). The CO 2 concentration of jars headspace was measured on the days of 1, 3, 5, 7, 12 and 17 after substrate addition. According to the CO 2 emission rate, the stable isotope 13 C of respired CO 2 was measured on Day 12 after substrate addition when microbial growth was stable. The CO 2 emission rate and the CO 2 stable isotope 13 C were also measured before substrate addition with a GasBench II system coupled to a Delta V Advantage IRMS (Thermo Scientific). Soil organic carbon (SOC) derived CO 2 (Priming effect) was calculated using equations1 and 2: where δ 13 C R is the δ 13 C of CO 2 from soil emission, δ 13 C so c and δ 13 C sub are the δ 13 C of soil and added substrates, and f som and f sub are the contribution of soil and added substrates to the total amount of released CO 2 56 . S oil characteristics a nalysis . The concentration of Fe bound organic carbon (Fe-OC) was quantitatively measured on the basis of the citrate-bicarbonate-dithionite (CBD) method 57 . Briefly, in a reduction treatment, a solution (15 ml, pH=7) containing 0.11 M sodium bicarbonate and 0.27 M trisodium citrate was added into 0.25 g freeze-dried soil and vortex oscillation in an 80°C environment heated by water bathing for 15 min. A reducing agent, sodium dithionite, was then added into mixture and 80°C water bathing for another 15 min. In the control treatment, instead of CBD extraction, soil was extracted with sodium chloride (NaCl) at an equivalent ionic strength. After centrifuging and filtering, the residual particles of both reduction and control treatments were rinsed with 5 ml DI water four times, freeze-dried, grounded to <100 μm and HCl-fumigated to remove the inorganic carbon, then were measured to get the SOC content. Fe-bound SOC content was calculated with the following equation: The concentration of water-soluble phenolics in soil was estimated using the Folin-Ciocalteu method with p -hydroxybenzoic acid as standard. Soil (5 g) and deionized water (25 ml) were shaken at a frequency of 200 rpm for 20 hours. After filtering through a 0.45 μm filter and diluting 1:5, the extracted solution (1 ml) was mixed with 2 ml Na 2 CO 3 -NaHCO 3 buffer (pH 10), 1 ml Folin-Ciocalteu reagent, and 1 ml 0.4 mol l -1 Na 2 CO 3 ,and kept in dark at 37°C for 15 min. Finally, the absorbance was measured at a wavelength of 750 nm using a spectrophotometer (Thermo Spectronic 200, USA). Standard phenolic absorbance was also measured with the same procedure. The concentration of water-soluble phenolics (measured in mg g -1 soil) was calculated using absorbance values of the standard and the soil samples 58 . Microb ial analysis. Microbial biomass carbon (MBC) concentration was measured using chloroform fumigation technique 59 . DNA of three layers soil was extracted using a MoBio PowerSoil Kit (MoBio Laboratories, Carlsbad, CA, USA) following the manufacturer’s instructions. The V4 region of bacterial 16S rRNA gene and the ITS1 region of fungi gene were amplified during PCR to characterize bacterial and fungal communities, respectively 60,61 . Quantitative PCR (qPCR) was performed to estimate the abundance of bacteria and fungi of soil 62 . The sequence data of 16S and ITS were retrieved from another paper by our team (deposited in the NCBI Sequence Read Archive (SRA) database under accession number SRP211776). Enzyme activity . Two oxidase enzymes (phenol oxidase and peroxidase) were assayed using the substrate of L-DOPA and two hydrolase were measured using the respective substrates of 4-MUB-β-D-glucopyranoside and 4-MUB-β-D-cellobioside. Soil (1.25 g) was added to 50 mM acetate buffer (125 ml, pH 5.0) and shaken for 5 min at 200 rpm to make a suspension 63,64 for following analysis. Two hydrolase of β-1,4-glucosidase and cellobiohydrolase were detected fluorometrically in the microplates. After filtering, 200 μl of sample solution was added to sample control, sample assay and quench tubes, while 200 μl of acetate buffer was added to blank, reference, and negative control tubes. Acetate buffer (50 μl) was also added to blank and sample controls, while 50 μl of 10 μM 4-methylumbelliferone substrate was added to reference and quench tubes, or 50 μl of 4-MUB-conjugated substrates added to negative control and sample assay tubes. All microplates were incubated at 20°C in the dark for four hours before the addition of 10 μl of 1.0 M NaOH to stop the reaction. Fluorescence was measured using a microplate fluorometer with 365 nm excitation and 450 nm emission filters. The fluorescence of the negative controls and the quenching were measured together and expressed in nmol h -1 g -1 . Two oxygen enzyme activities were measured spectrophotometrically. Similar to the fluorometric analysis, 200 μl of filtered soil solution was added to tubes of sample control and sample assay; 200 μl of acetate buffer was added to tubes of blank and negative controls; 50 μl of acetate buffer was add to tubes of blank and sample controls; and 50 μl of 25 mmol L-DOPA was added to tubes of negative control and sample assay. For peroxidase assays, another 10 μl of 0.3% H 2 O 2 was added to each tube. All microplates were incubated at 20°C in dark for 20 hours. The absorbance was measured at 450 nm using a microplate spectrophotometer (Varioskan Flash, Thermo, USA). The activity was expressed in μmol h -1 g -1 . Data analysis . Differences in soil biogeochemical characteristics and soil CO 2 emission across soil layers were assessed for significance using ANOVA with Duncan test. ANOVA was also used to assess the differences of soil CO 2 emission when three substrates were added. t - test was used to assess the differences in soil CO 2 emission between 8°C and 18°C. All statistical analyses were conducted in SPSS 20.0 for Windows (SPSS, Chicago, IL, USA) and all figures were created in Origin 8.0. Microbial raw sequence data was processed using QIIME2 (version 2018.11) 65 and diversity was performed using R (Version 3.3.1). Piecewise structure equation model (SEM) was used to compare the direct and indirect effects of temperature, inputting substrate, soil microbes (MBC), enzymes, soil phenolics /nitrogen nutrient on soil CO 2 emission of each layer, using piecesiseSEMpackage in R. One specific hypothesized interactions included in the model was outlined in Supplementary Fig. 12. The temperature and inputting fresh substrate were specified as exogenous variables; soil enzymes, MBC, soil carbon components, and nitrogen nutrient were endogenous variables. Paths were included from temperature and inputting fresh substrate to all endogenous variables, then to soil CO 2 emission. The model was used in linear mixed effects models with block as a random effect in the piecewise SEM. A stepwise removal of the least significant paths from the models was conducted until either the AIC did not decrease with removal of the next parameter or there were no remaining parameters with p > 0.1. Results Soil properties. Of the soil carbon quality proxies, the concentration of Fe-OC was significantly higher at ETzone than at aerobic surface and anaerobic deep soil ( p < 0.01), reflecting the abundance of recalcitrant and aggregated protected carbon in ETzone. H 2 O-PhO was higher at ETzone and anaerobic deep soil than at aerobic surface soil ( p < 0.01; Table 1). In contrast with soil carbon quality proxies, the microbial activities including soil MBC ( p < 0.01), the Shannon diversity of bacteria ( p 0.05), were significantly lower at ETzone than at aerobic surface and anaerobic deep soil. The qPCR results showed that bacterial ( p 0.05) copy numbers were lower in anaerobic deep soil than in aerobic surface soil. In terms of soil enzymes, phenolic oxidase ( p > 0.05) and β-1-4-glucosidase ( p > 0.05) had the highest activities at aerobic surface soil, while the highest activity of peroxidase ( p < 0.05) and cellobiohydrolase ( p < 0.01) were at ETzone (Table 1). Response to warming . After a long-term incubation, the potential CO 2 emission rate at ETzone (28.31 ± 3.55 μg CO 2 g -1 d -1 ) was significantly lower than the other two layers (surface: 48.44 ± 3.59 μg CO 2 g -1 d -1 ; subsurface: 38.06 ± 1.71 μg CO 2 g -1 d -1 ; p 0.05) in warming treatment. Different from the ETzone, the CO 2 emission in aerobic surface and anaerobic deep soil were significantly enhanced by warming ( p < 0.01 for both; Fig 2a), with Q 10 of 2.14 and 1.51, respectively (Table 1). These results indicated that the soil at ETzone was not sensitive to warming. Warming also had significantly positive effect on MBC concentration of all three layers soil (Supplementary Fig. 4). With soil degrading from field anaerobic conditions to aerobic scenario, the warming response of CO 2 emission of both ETzone and anaerobic deep soil were significantly enhanced ( p < 0.01 for both; Fig 2a), with increments of 65.7% to 83.4% at ETzone (Supplementary Fig. 5a) and 68.4% to 89.2% at anaerobic deep soil (Supplementary Fig. 5b). In future aerobic scenario, the activity of soil oxidase enzymes including phenol oxidase and peroxidase were significantly higher than in field conditions and further increased by warming (Supplementary Fig. 6). From field anaerobic conditions to future degraded scenario, the increment of CO 2 emission facilitated by warming of the upper 100 cm soil profile increased by 33.7% (ranging from 84.62 ± 3.67 μg CO 2 g -1 d -1 to 127.92 ± 7.38 μg CO 2 g -1 d -1 ; Supplementary Fig. 7). Response to p riming . The response to priming of soil carbon emission varied with soil conditions and layers. In field conditions, new substrate addition increased soil CO 2 emission in all three layers, but significantly only at ETzone ( p 0.05 for both), demonstrating that the ETzone was more sensitive to new substrate than the other two layers. In future degraded aerobic scenario, the increments of CO 2 emission caused by substrate addition were significant at both ETzone and anaerobic deep soil ( p < 0.01 for both; Fig 2b). Substrates like glucose and cinnamic acid also enriched fungal abundance (Supplementary Fig. 8), increased phenol oxidase and peroxidase activities significantly in both field conditions and future degraded scenario (Supplementary Fig. 9). The priming response of soil carbon at each layer varied with substrates and conditions. In the field conditions, CO 2 emission was significantly enhanced by oxalate in the aerobic surface soil, by cinnamic acid in the anaerobic deep soil, and by both in the ETzone. Glucose did not show any significant influence on soil CO 2 emission in any layers (Fig 3a). In the degraded aerobic scenario, however, every substrate increased soil CO 2 emission of all three layers (Fig 3b). Comparison of priming and warming . The log values of the increased CO 2 emission caused by priming and warming were compared in three layers, to predict the main factor influencing soil CO 2 emission. In field conditions, we found that warming had a higher log value than priming at aerobic surface and anaerobic deep soil. However, at ETzone, the log value of priming was higher than warming. These results indicated that aerobic surface and anaerobic deep soil were more influenced by warming and the ETzone was more influenced by priming. In the future degraded scenario, the log value of both warming and priming increased, with different increments at ETzone and anaerobic deep soil (Fig 4). Similar increment of warming and priming at anaerobic deep lead to the main influencing factor still was warming. At ETzone, greater increment of warming resulted in the main influecning factor became wamring (Fig 4). In terms of different substrates, each showed different priming log value for three layers, substrate with strong priming effect (Fig 3) had higher log value (Supplementary Fig. 10). Determinants of CO 2 emission. Soil variables varied significantly in warming and priming responses. The significantly correlated soil variables were differed among three layers (Supplementary Fig. 11). SEM results showed that soil variables could explain 75%, 75% and 61.1% of the variability of CO 2 emission of aerobic surface, ETzone and anaerobic deep soil, respectively (Fig 5). At aerobic surface soil, NH+ 4 ( p < 0.01), MBC ( p 0.05) had positive effect on CO 2 emission. NH+ 4 also indirectly affect CO 2 emission through being negatively correlated with MBC ( p > 0.05) and positively correlated with β-1-4-glucosidase ( p < 0.01). Temperature and inputting substrate exerted strong indirect effect on CO 2 emission through their positive correlation with NH+ 4 ( p < 0.001) and MBC ( p < 0.01; Fig 5a). For anaerobic deep soil, cellobiohydrolase ( p 0.05) had directly positive effect on CO 2 emission, and H 2 O-PhO ( p > 0.05) had negative effect on CO 2 emission. Temperature exerted strong indirect effects on CO 2 emission through its positive correlation with MBC ( p < 0.01), and the inputting substrate exerted indirect effect on CO 2 emission through its positive correlation with cellobiohydrolase ( p < 0.05) and negative correlation with MBC ( p < 0.01) and H 2 O-PhO ( p < 0.05; Fig 5c). At ETzone, CO 2 emission was positively affected by soil phenolic oxidase ( p 0.05), but negatively by H 2 O-PhO ( p > 0.05). MBC also indirectly affected CO 2 emission through being negatively correlated with H 2 O-PhO ( p > 0.05). Temperature and inputting substrate exerted strong indirect effect on CO 2 emission through the positive correlation with MBC ( p < 0.001), phenolic oxidase ( p < 0.01) and H 2 O-PhO ( p < 0.01; Fig 5b). Taken together, the most important drivers of CO 2 emission were temperature ( p < 0.001) and NH+ 4( p < 0.01) at aerobic surface soil, and were temperature, the inputting substrate and cellobiohydrolase ( p < 0.01 for all) at anaerobic deep soil. The main drivers of CO 2 emission at ETzone were inputting substrate ( p < 0.001) and phenolic oxidase ( p < 0.01; Fig 5). Discussion The CO 2 emission at ETzone was significantly lower than the upper aerobic surface soil (41.6%) and anaerobic deep soil (34.4%). Moreover, 10°C warming failed to stimulate ETzone CO 2 emission significantly, while significantly stimulated that in the upper aerobic surface and anaerobic deep soil in the conditions of pristine field. In contrast with most other findings labeling ETzone as a hotspot in the ecosystem 1,4,5,34 , the lowest CO 2 emission rate and least warming response of the ETzone at the subsurface of peatland in our study showed that it was inert and resistant to warming. Such phenomenon can be explained by the constitution and characteristics of the carbon at ETzone soil. It is known that the ETzone in peatland experienced most severe disturbance during long-term water table fluctuation, which consumes great amounts of labile carbon and enriches abundance of Fe-OC (Table 1). Fe-OC is recalcitrant to soil microbes and warming resistant 36 . Higher CO 2 emission from the aerobic surface and the anaerobic deep soil, consistent with other studies 13,14 , reflects the presence of adequate plant substrates accessible to microbes 35 and high microbial activity 36 (Table 1, Supplementary Fig. 4). The relatively lower Fe-OC likely contributed to the higher sensitivity of the anaerobic deep soil to warming 37 (Table 1). The inert feature and non-responsive characteristic make ETzone a buffer against soil carbon loss from peatlands under warming climate, rather than a hotspot. However, the warming resistant characteristics of the ETzone was compromised when exposed to aerobic conditions simulating future degraded scenario. Our results showed that the CO 2 emission from the ETzone became significantly sensitive to warming, and the deeper adjacent soil became even more sensitive to warming under aerobic conditions. These results were consistent with a previous study indicating that drier or aerobic subsurface soil was sensitive to warming 10 . In our study, the presence of O 2 increased the activity of the oxidase enzyme (Supplementary Fig. 6), which enhanced soil carbon decomposition 38 . Compared to that of pristine field conditions, the increment of CO 2 emission caused by warming of the upper 100 cm soil profile was 33.7% higher in the degraded scenario. Therefore, about one third more CO 2 will be stabilized by the protection of the inert and warming-resistant ETzone, given it being kept in an anaerobic dominated condition (Supplementary Fig. 7). Noteworthy, contrary to the warming response, CO 2 emission at the ETzone was significantly sensitive to priming of fresh substrates, even under anaerobic conditions, no matter labile carbon of oxalate and glucose, or recalcitrant carbon of cinnamic acid. There are some possible reasons for this phenomenon. Due to frequent water table fluctuation, SOC at ETzone was dominated in mineral associations (Table 1) and the microbes were limited by energy 33 . Exo-substrates representing both labile and recalcitrant carbon alleviated energy limitation of the microbes, especially for fungi 33 which were of relatively low abundance in original soils (Table 1) and increased greatly in diversity and relative abundance after substrate addition (Supplementary Fig. 8). Fungi could excrete substantial extracellular enzymes (Supplementary Fig. 9a and c) to decompose recalcitrant carbon and break organo-mineral complexes, liberating organic compounds from protected associations or aggregates 39,40 for further decomposition. Different from that of ETzone, CO 2 emission from the aerobic surface soil remained stable after substrate addition, except with oxalic acid (Fig 3). With strong metal-complexing ability, oxalic acid can cause strong priming effect by breaking mineral-organic associations, but is of limited bioenergetics use to microbes 39 . The significant influence of oxalic acid demonstrated that the surface soil was not energy-limited. In the anaerobic deep soil, CO 2 emission was also enhanced by the addition of substrates, although not significantly (Fig 2b), except for the addition of cinnamic acid (Fig 3), which contains an aromatic loop and is a complex product of lignin depolymerization 32 . The significant increase in soil CO 2 emission caused by cinnamic acid suggested that the deep soil was rich in recalcitrant carbon in the form of aggregates 36 . The protection offered by aggregates rendered the soil insensitive to other substrates. Combined with aerobic conditions which opened the enzyme “latch” 38 (Supplementary Fig. 9b and d), the priming effects of all substrates were enhanced in the degraded scenario, leading to further decomposition of liberated organic compounds (Fig 3b). We found that the response of the ETzone soil to warming and priming significantly differed from the other two layers. Under the field conditions, the aerobic surface soil was influenced more intensively by warming than by priming (Fig 4), which probably reflected that the activity of soil microbes and enzymes were suppressed by low temperature 36 . Warming could enhance soil carbon decomposition by favoring soil microbes and enzyme activity 36,41 . Similarly, the anaerobic deep soil was also influenced more intensively by warming than priming, though with a weaker response to warming than the surface soil (Fig 4). This is ascribed to the long-term anaerobic conditions and the limited soil nutrients in the deep soil 42 (Table 1). Even when soil conditions changed from the field anaerobic to the future degraded aerobic, the dominant effect at anaerobic deep soil kept as warming. With the strongest priming effect among the three layers, the dominant effect for ETzone was priming. Soil carbon in the ETzone was substantially stabilized due to the abundant recalcitrant carbon inaccessible for microbes, e.g., Fe-OC 36 (Table 1), and the microbes were limited in nutruent. Substrate addition helped mitigate this limitation by supplying carbon and energy to soil microbes 43 . When soil conditions changed from the field anaerobic to the future degraded aerobic, however, the dominant effect for ETzone changed from priming to warming (Fig 4). In oxygen unlimited environment, aerobic decomposition could releases nutrient from soil aggregates and recalcitrant carbon, easing the limitation of energy and nutrient on soil microbes. Thus, even with increase of effect by both warming and priming, priming was no longer the dominant factor. The main determinants of soil CO 2 emission at ETzone also differed with the other two layers. Our result showed that aerobic surface soil CO 2 emission was significantly determined by NH+ 4, demonstrating that surface soil was in N-limited conditions, consisting with previous researches which showed that peatlands on Zoige was a nitrogen limited system 44 . As with observed in warming response, temperature was indeed one of critical factors in determining aerobic surface soil CO 2 emission, through indirectly influencing soil microbial biomass 45 (Fig 5). For CO 2 emission from anaerobic deep soil, temperature was a critical factor too 15 , since warming could increase soil microbial biomass and cellobiohydrolase, which are limited by temperature at both surface and deep soil 45,46 . Soil microbes at these two layers might also be in energy limited conditions, for their activities increased significantly after fresh substrate addition 33 . The significant direct influence of cellobiohydrolase at anaerobic deep soil demonstrated that the carbon in this layer was more complex than in aerobic surface soil 36 . Contrasted with that at aerobic surface and anaerobic deep soil, the CO 2 emission at ETzone was mainly determined by phenolic oxidase (Fig 5), indicating that substantial recalcitrant carbon subject to phenolic oxidase existed at ETzone, and that the released CO 2 may mainly result from the decomposition of these recalcitrant carbon. Phenolic oxidase and microbes at ETzone were limited in energy and nutrient, and stimulated by fresh carbon. The limitation of soil phenolic oxidase and microbe at ETzone could better explain the significant response of ETzone CO 2 emission to added substrates. From the CO 2 emission characteristics and its major determinants, we can see that the ETzone of the subsurface soil was characterized with both warming-resistance and substrate-sensitivity (Fig 6). The warming-resistant ETzone could serve as a buffer to prevent depletion of carbon stock inside and below peat due to decomposition in a warming climate. For example, for northern peatlands with a carbon stock of 547 Pg (an average depth of 230 cm) 47,48 , and an ETzone presumably of 23.3 cm below the peatland surface (Fig 1), the inert ETzone would protect about 89.9% of the carbon stored inside and below it from warming-induced decomposition. However, due to the priming-sensitiveness of the ETzone, such protection can easily collapse through priming. Thus, a non-priming ETzone will be beneficial to maintain carbon stability of peatlands. A community with stable plant composition produces relatively constant amounts and type of root exudates 49,50 , and stable hydrological conditions are helpful for minimizing substrate exchange among soil layers 51 and the positive priming effects on soil carbon, ultimately decreases carbon loss 52 . With ever more severe disturbance of climate change and human activities, carbon loss from peatlands had substantially increased in the last few decades 53,54 . Several mechanisms were proposed as the key to keep peatlands carbon stable, e.g., the enzyme “latch” theory that emphasizes the importance of anaerobic environment 38 in limiting enzyme activities and suggests sustaining a stable and high water table to protect carbon stock; and the “iron gate” theory about the essential role of Fe-OC to decrease carbon decomposition and protection of soil aggregation 40,55 . In our study, we found that maintaining a relatively constant status of the ETzone at the subsurface soil is critical in protecting peatland carbon stock. Because of the priming-sensitiveness of ETzone, the priority is to establish a non-priming ETzone by maintaining intact vegetation composition to reduce the possibility of intensified positive priming effect; Furthermore, due to alleviation in warming-resistance of the ETzone soil in the degraded scenario, another important step is to sustain the depth of warming resistant ETzone through maintaining a constant water regime (Fig 6). Our results also illustrated that ETzones along environmental gradients will not always be “hotspot” of biogeochemical processes and interactions, but could be a “buffer” against carbon loss. However, whether this theory is applicable across all terrestrial ecosystems needs further investigation. Considering the limitation of the incubation experiment that missed the information of seasonal variations in temperature, phenology and soil moisture, there was still much uncertainty in illuminating in situ carbon dynamic. Our results provided a compelling evidence of the stabilizing function of the ETzone on peatland carbon stock. Previous studies that had highlighted the critical role of subsoil in peatlands carbon dynamic, without considering the buffer capacity of the ETzone, may have overestimated the carbon loss under warming climate of peatlands, and underestimated the carbon loss through priming by shift in vegetation composition under climate change and human disturbance. Moreover, including the function of the ETzone in ecological modeling will be beneficial to accurately simulate the impacts of global change on SOC dynamic of peatlands. With climate change and the induced deeper plant root distributing, the stabilization of the ETzone would be non-negligible in predicting peatlands carbon dynamics. Declarations Acknowledgments We appreciate the assistance in language editing by Ms. Wan Xiong. This work was supported by the Strategic Priority Research Program of Chinese Academy of Science (XDA200500404), the Second TIBETAN Plateau Scientific Expedition (2019QZKK0304) and Key Research Program of Frontier Sciences, CAS, QYZDB-SSW-DQC007. Author contributions : H.C, L.L, N.W, H.W, and Y.W designed the experiments. L.L, J.T, and D.X performed the experiment. L.L and H.C analyzed the data. L.L, H.C, Y.W, N.W, X.X, H.W, C. P, and M.W wrote the manuscript. Competing interests: The authors declare no competing interests. Additional information Supplementary Materials: Supplementary Figures. 1 to 12 Correspondence and requests for materials should be addressed to N.W and Y.W. References 1 Juutinen, S. et al. Major implication of the littoral zone for methane release from boreal lakes. Global Biogeochemical Cycles 17 , doi:10.1029/2003GB002105, doi:10.1029/2003gb002105 (2003). 2 Pegoraro, E. et al. 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Aerobic ETzone Anaerobic Soil characteristics Fe-OC (%) 3.65 ± 0.11B 5.36 ± 0.68A 2.75 ± 0.06B H 2 O-PhO (mg kg -1 ) 191.9 ± 6.39B 357.76 ± 13.199A 382.26 ± 34.37A Decompos er MBC (mg kg -1 ) 3119.1 ± 256.78A 289.89 ± 33.6C 1262.37 ± 111.33B Bacteria Shannon diversity [ H 、 ] 5.59 ± 0.15A 4.33 ± 0.17C 5.07 ± 0.11B Bacteria numbers (x10 6 ) 142.29 ± 21.31A 29.46 ± 5.21B 20.97 ± 3.2B Fungi Shannon diversity [ H 、 ] 2.22 ± 0.03 1.95 ± 0.13 2.09 ± 0.01 Fungi numbers (x10 6 ) 3.79 ± 2.37 0.13 ± 0.07 0.17 ± 0.11 Phenolic oxidase (μmol h -1 g -1 ) 5.8 ± 0.56 4.87 ± 0.3 3.93 ± 0.72 Peroxidase (μmol h -1 g -1 ) 3.08 ± 0.41ab 3.8 ± 0.07a 2.51 ± 0.08b β-1-4-glucosidase (nmol h -1 g -1 ) 5.5 ± 0.14 4.97 ± 0.6 3.61 ± 0.59 Cellobiohydrolase (nmol h -1 g -1 ) 57.81 ± 6.15A 71.38 ± 9.33A 9.27 ± 2.24B Carbon emission CO 2 rate (μg CO 2 g -1 d -1 ) 48.44 ± 3.59A 28.31 ± 3.55B 38.06 ± 1.71A Q 10 2.14 ± 0.13A 1.37 ± 0.13B 1.51 ± 0.08B Values are mean ± SE (n = 3). Fe-OC: Fe-bound soil organic carbon; H 2 O-PhO: Water-soluble phenolics; MBC: microbial biomass carbon. Letters indicated the significant difference of soil properties among three layers (Duncan test; p < 0.05 for lowercase and p < 0.01 for uppercase). Additional Declarations There is NO Competing Interest. Supplementary Files Supplementary.docx Supplementary Materials: Supplementary Figures. 1 to 12 Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-849966","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":50418602,"identity":"869e2f1e-b543-4567-9838-76d2c0ee689b","order_by":0,"name":"Liangfeng Liu","email":"","orcid":"","institution":"Chengdu Institute of Biology, Chinese Academy of Science","correspondingAuthor":false,"prefix":"","firstName":"Liangfeng","middleName":"","lastName":"Liu","suffix":""},{"id":50418603,"identity":"ed54c2fd-a9cb-4a58-ad7a-19e8b1e91854","order_by":1,"name":"Huai Chen","email":"","orcid":"","institution":"Chengdu Institute of 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06:21:02","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-849966/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-849966/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":13198544,"identity":"24c5155e-0055-40b5-bf47-6ec38da28ad3","added_by":"auto","created_at":"2021-09-08 21:41:40","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":936605,"visible":true,"origin":"","legend":"CO2 emission rate of global peatlands soil from ETzone and the other two zones (dot figures in the right) which was divided according to the water table of global peatlands (box figure in the left).","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/a07b752bde78b56ab62048b8.png"},{"id":13199018,"identity":"243342af-ceb4-4045-bc0c-d2a376030a5e","added_by":"auto","created_at":"2021-09-08 21:47:40","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":272988,"visible":true,"origin":"","legend":"CO2 emission rate from environment transitional zone (ETzone) and two adjacent layers under the effect of (a) warming and (b) substrate priming effect. Filled symbol and orange label: field conditions; Hollow symbol and grey label: future degraded scenario. Values are mean ± standard error (n=3) (Duncan test, *: p \u003c 0.05, **: p \u003c 0.01).","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/67b8a730fe7546bfb7c28192.png"},{"id":13198740,"identity":"88915851-3203-4038-a9e0-3d8d863f9684","added_by":"auto","created_at":"2021-09-08 21:44:40","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":287158,"visible":true,"origin":"","legend":"Priming effect of three substrates on three layers soil, under the conditions of (a) field and (b) future degraded scenario. Value ± SE, n = 3. (t test, **: p \u003c 0.01, *: p \u003c 0.05, NS: p \u003e 0.05).","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/490047ddb24861c5521c6541.png"},{"id":13198545,"identity":"b22ae229-4e75-4720-8edc-5e893062869b","added_by":"auto","created_at":"2021-09-08 21:41:40","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":278137,"visible":true,"origin":"","legend":"Ratios of the log value of the increased CO2 emission in the conditions of priming to warming of three layers soil. At the environment transitional zone (ETzone: Subsurface) and anaerobic deep soil, the increase of symbol size indicates the gradual change of soil conditions from anaerobic to aerobic (Values are means of ± standard error, n =3). ","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/223c99d4280c16bb070206c7.png"},{"id":13198549,"identity":"0ed1b4b4-4bbb-4790-bc56-3bb5c02ac141","added_by":"auto","created_at":"2021-09-08 21:41:40","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":3621176,"visible":true,"origin":"","legend":"The main drivers of soil CO2 emission of (a) aerobic surface, (b) ETzone and (c) anaerobic deep soil. Glu-ase: β-1-4-glucosidase; H2O-PhO: Water-soluble phenolics; Phe-ase: Phenolic oxidase; Cell-ase: Cellobiohydrolase. Blue and orange arrows indicate positive and negative relationships, respectively. Number adjacent to arrows are the standardized path coefficients (*: p \u003c 0.05; **: p \u003c 0.01; ***: p \u003c 0.001). The conditional r2 (based on the variance of both the fixed and random effect) for the component mixed effects of both the models are shown in the boxes for the response variables.","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/d1333d6117049861df83a337.png"},{"id":13198550,"identity":"c9431bf6-4c8d-4f1d-ae12-d7914e433f67","added_by":"auto","created_at":"2021-09-08 21:41:40","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":863996,"visible":true,"origin":"","legend":"The role of ETzone of subsurface soil in peatlands. In changing climate, two steps were crucial in stopping carbon loss: 1) easing priming effect by maintaining intact vegetation and root exudate; 2) sustaining relative high and constant water regime zone against warming on ETzone.","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/9c0b3c673a6fe69382a4add9.png"},{"id":19963790,"identity":"017f1feb-f4e5-4a92-aa16-2ac1c541d35e","added_by":"auto","created_at":"2022-04-05 08:26:13","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2373371,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/9738f8ea-ddd1-4e08-ae98-f048a767ca44.pdf"},{"id":13198546,"identity":"82ba8d0a-89f9-4b96-a979-afd98d5d3a1f","added_by":"auto","created_at":"2021-09-08 21:41:40","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":1301915,"visible":true,"origin":"","legend":"Supplementary Materials: Supplementary Figures. 1 to 12","description":"","filename":"Supplementary.docx","url":"https://assets-eu.researchsquare.com/files/rs-849966/v1/0846e81b9187798a83bf0d65.docx"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e Competing Interest.","formattedTitle":"\u003cp\u003eAn Environment Transitional Zone Buffers Peatlands Carbon Loss\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAn environment transitional zone (ETzone), which connects zones with totally different environment and biogeochemical features,\u0026nbsp;exists in\u0026nbsp;the interfaces\u0026nbsp;along an environmental gradient, e.g.,\u0026nbsp;the littoral zone between aquatic and terrestrial\u003csup\u003e1\u003c/sup\u003e, or\u0026nbsp;the active freeze-thaw layer\u0026nbsp;in\u0026nbsp;discontinuous\u0026nbsp;permafrost\u003csup\u003e2\u003c/sup\u003e,\u0026nbsp;or the redox interface along the soil profile of peatlands\u003csup\u003e3\u003c/sup\u003e. Being in the interface of two adjacent zones, the ETzone possesses the characteristics of both and is often regarded as a hotspot of biogeochemical processes\u003csup\u003e1\u003c/sup\u003e, e.g., the littoral zone is hotspots of CH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e1\u003c/sup\u003e and CO\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e4\u003c/sup\u003eemission; a temperate forest transitional slopes is hotspots of CO\u003csub\u003e2\u003c/sub\u003e emission and CH\u003csub\u003e4\u003c/sub\u003e uptake\u003csup\u003e5\u003c/sup\u003e. In addition, owing to frequent disturbances, ETzones are also regarded as the most vulnerable part of ecosystems\u003csup\u003e6\u003c/sup\u003e, especially for those fragile ecosystems, such as peatlands.\u003c/p\u003e\n\u003cp\u003eIn peatlands, an ETzone develops during water table fluctuation\u003csup\u003e7\u003c/sup\u003e (Supplementary Fig. 1), which is at the subsurface of soil profile and characterized by alternatively aerobic and anaerobic environment. Above the ETzone is aerobic surface soil under permanent aerobic conditions and abundant in living plants and roots (i.e., acrotelm); below the ETzone is anaerobic deep soil perpetually water logged and entirely without\u0026nbsp;fresh plant roots (i.e., catotelm)\u003csup\u003e8,9\u003c/sup\u003e. Adjacent to the two soil layers, the ETzone acquires most severe disturbance due to the long term fluctuation of water table, resulting in the soil biogeochemical characteristics quite different from adjacent soil\u003csup\u003e3,7\u003c/sup\u003e (Table 1). However, whether this ETzone is a hotspot in soil carbon loss remains uncertain, especially under the situation of intensified climate warming and priming\u003csup\u003e10-12\u003c/sup\u003e.\u003c/p\u003e\n\u003cp\u003eIntensified climate warming is expected to increase soil microbial decomposition and carbon export, influencing peatlands soil carbon dynamics\u003csup\u003e13,14\u003c/sup\u003e. A non-disturbing\u003cem\u003e\u0026nbsp;in situ\u0026nbsp;\u003c/em\u003eexperiment showed that 1\u0026deg;C\u0026nbsp;warming could accelerate ecosystem respiration by 52%-60%, in which at least 69% increase originated from the soil at 25-50 cm\u003csup\u003e15\u003c/sup\u003e. Other \u003cem\u003ein situ\u0026nbsp;\u003c/em\u003ewhole soil profile warming experiments verified that subsoil is vulnerable to warming\u003csup\u003e16,17\u003c/sup\u003e.\u0026nbsp;A lab simulating experiment\u0026nbsp;showed that subsoil (\u0026gt; 20 cm) was responsible for 65%-70% increase of whole profile soil carbon emission caused by warming\u003csup\u003e18\u003c/sup\u003e. We further explored the response of soil carbon to warming of global peatlands from ETzone and the other two zones through compiling the data of CO\u003csub\u003e2\u003c/sub\u003e emission rate of global peatlands soil conducting by incubation under differed temperatures and ranging a depth of 0-10 cm to 100-120 cm (Supplementary Fig. 2). The three zones were defined according to the water table data of global peatlands which were also collected from vast publications (Supplementary Fig. 3). Despite the reactive characteristic of subsoil in \u003cem\u003ein situ\u0026nbsp;\u003c/em\u003eand lab experiments, we failed to observe the anticipated higher warming response and Q\u003csub\u003e10\u003c/sub\u003e for ETzone in the compiled data (Fig 1). Thus, by treating the peat profile as a whole and not considering the peculiarity of the ETzone at the subsurface layer\u003csup\u003e13,19\u003c/sup\u003e, these experiments might have reached to inconclusive results, though established an active role of subsoil of peatlands in projecting soil carbon feedbacks to climate warming\u003csup\u003e10,20\u003c/sup\u003e. More research is needed to understand the characteristics of the ETzone of peatlands and its response to climate warming in terms of soil carbon stability.\u003c/p\u003e\n\u003cp\u003ePriming is a composite effect of enhanced soil microbial growth and activity by increased carbon availability from vegetation roots/litters, altered pH values, soil aggregation and microbial community composition\u003csup\u003e21-23\u003c/sup\u003e. Researches showed that soil organic carbon (SOC) respiration can be 380% to 1200% higher with the input of plant originating compounds\u003csup\u003e24\u003c/sup\u003e, and that such priming effect could persist over long time\u003csup\u003e25\u003c/sup\u003e. In peatlands, the soil is found more vulnerable to the priming effect\u003csup\u003e26,27\u003c/sup\u003e and the priming effect on SOC was even strengthened under intensified climate change and human activities (e.g., water table drawdown, over-grazing)\u003csup\u003e28,29\u003c/sup\u003e. Similar with the threat from warming, there is so far no data for the effect of priming on SOC stability at ETzone, despite the priming effect on soil carbon stability has been widely concerned\u003csup\u003e22,30,31\u003c/sup\u003e.\u003c/p\u003e\n\u003cp\u003eIn this study, in order to understand the carbon dynamics of ETzone under intensified warming and priming, the soil CO\u003csub\u003e2\u003c/sub\u003e emissions and the determinants of ETzone were quantified under simulating conditions on Zoige plateau, which is a typical high-altitude minerotrophic peatlands distributed on the eastern edge of the Qinghai-Tibetan Plateau and with the global importance in predicting climate. The ETzone in our peatland was classified according to the long-term water table fluctuation (Supplementary Fig. 1). Soil samples from the ETzone (subsurface: 26-75 cm) and the two adjacent layers (aerobic surface layer: 0-25 cm; anaerobic deep layer: 76-100 cm) were incubated under the\u0026nbsp;pristine\u0026nbsp;field conditions (surface, aerobic; subsurface, anaerobic; deep, anaerobic) with three treatments:\u0026nbsp;control (mean temperature in growing season:\u0026nbsp;8\u0026deg;C), warming (18\u0026deg;C) and\u0026nbsp;substrate addition (8\u0026deg;C). Three substrates (oxalic acid, glucose, and cinnamic acid) found primarily in plant litter/roots exudate\u003csup\u003e32\u003c/sup\u003e were added into soils to simulate the priming effect. The same treatments were also applied to soil samples that were incubated under aerobic conditions to simulate the future degraded scenario when the water table in peatlands decreases severely and the peat soil of both ETzone and the deep layer are exposed in aerobic environment. The CO\u003csub\u003e2\u003c/sub\u003e emission rate was monitored weekly to quantify the response of soil carbon to warming. The \u0026delta;\u003csup\u003e13\u003c/sup\u003eC of respired CO\u003csub\u003e2\u003c/sub\u003e was analyzed before and after substrate addition to quantify the priming effect of three substrates on soil carbon stability\u003csup\u003e33\u003c/sup\u003e.\u0026nbsp;Soil variables of carbon component\u0026nbsp;(Fe bound organic carbon, Fe-OC;\u0026nbsp;water-soluble phenolics: H\u003csub\u003e2\u003c/sub\u003eO-PhO), nitrogen nutrient (NH+ 4), microbial indexes (microbial biomass carbon: MBC; relative abundance), and enzyme activities\u0026nbsp;(phenolic oxidase, peroxidase,\u0026nbsp;\u0026beta;-1-4-glucosidase, and cellobiohydrolase)\u0026nbsp;representing carbon quality, soil nutrient, and decomposers were determined during incubation to explore the main determinants of soil CO\u003csub\u003e2\u003c/sub\u003e emission of three layers, by piecewise structure equation model (SEM).\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003e\u003cstrong\u003eSampling\u003c/strong\u003e\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eSoil samples were collected from Ruokeba peatland\u0026nbsp;on the Zoige Plateau, northeastern\u0026nbsp;Qinghai-Tibet Plateau.\u0026nbsp;Zoige\u0026nbsp;Plateau is characterized\u0026nbsp;with\u0026nbsp;low temperature (mean\u0026nbsp;annual temperature of -3.3\u0026deg;C)\u0026nbsp;and\u0026nbsp;rich river system\u0026nbsp;that\u0026nbsp;accelerates\u0026nbsp;peat\u0026nbsp;accumulation. Peatlands\u0026nbsp;on Zoige\u0026nbsp;Plateau covers\u0026nbsp;an area of 4,605 km\u003csup\u003e2\u003c/sup\u003e, with an average peat depth of 0.2-6.0 m. The major vegetation comprises of \u003cem\u003eEquisetum ramosissimun, Scirpus triqueter,\u0026nbsp;\u003c/em\u003eand\u003cem\u003e\u0026nbsp;Blysmus sinocompressus.\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eInfluenced by human disturbance and climate change, water table at Ruokeba peatland decreased severely and influenced soil carbon dynamic for years. During long-term water table fluctuation, a distinct environment transitional zone (ETzone) at subsurface soil (characterized with periodically aerobic and anaerobic due to water table fluctuation) was shaped out. Soil adjacent to ETzone includes upper aerobic surface soil (characterized with predominantly aerobic, abundant in living plants and roots) and lower anaerobic deep soil (characterized with persistent water logging and lacks of plant roots). Soil cores (105 cm in depth) were sampled at three randomly selected sites in May 2015, using a vibracore with a diameter of 8 cm. All soil cores were divided into three layers according to the water table fluctuation \u0026nbsp;(Supplementary Fig. 1): aerobic surface soil (0-25 cm), subsurface soil (ETzone: 26-75 cm) and anaerobic deep soil (76-100 cm). Each layer soil from the same site was mixed, sieved (2 mm) and transported to laboratory using sterile bags in ice boxes. In laboratory, soil sample was stored at 4\u0026deg;C for incubation and soil characteristics analysis, and at -20\u0026deg;C for microbial analysis. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eIncubation\u003c/strong\u003e\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eSoil was incubated in the conditions of field (surface, aerobic; subsurface, anaerobic; deep, anaerobic), future degraded scenario (aerobic), with treatments of control, warming and substrate addition.\u0026nbsp;Sieved soil (80 g)\u0026nbsp;of\u0026nbsp;each\u0026nbsp;layer\u0026nbsp;was placed in a 500-ml glass jar and sealed using a rubber stopper with two openings for\u0026nbsp;gas\u0026nbsp;sampling and headspace exchange.\u0026nbsp;Jars headspace was replaced\u0026nbsp;with N\u003csub\u003e2\u003c/sub\u003e or CO\u003csub\u003e2\u003c/sub\u003e-free air to create\u0026nbsp;anaerobic\u0026nbsp;and\u0026nbsp;aerobic\u0026nbsp;conditions, respectively\u003csup\u003e18\u003c/sup\u003e. In warming experiment, jars were placed at\u0026nbsp;8\u0026deg;C\u0026nbsp;(mean temperature in growing season) as control\u0026nbsp;and 18\u0026deg;C\u0026nbsp;as warming\u0026nbsp;(3 depths * 2 conditions * 2 temperatures * 3 replicates = 36 jars). After pre-incubation for seven days, the headspace gas was weekly sampled to measure\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission rate. At 12 hours before each sampling, the headspace was flushed with N\u003csub\u003e2\u003c/sub\u003e or CO\u003csub\u003e2\u003c/sub\u003e-free air again to\u0026nbsp;remove the accumulated CO\u003csub\u003e2\u003c/sub\u003e. The headspace gas was sampled by vacuum tube and CO\u003csub\u003e2\u003c/sub\u003e concentration was measured by gas chromatography (Agilent 7890A, Agilent Co., USA) with a flame ionization detector (FID) operating at 250\u0026deg;C. Soil water content was kept stable during incubation by constant weight and all jars were incubated for 154 days to evaluate long-term CO\u003csub\u003e2\u003c/sub\u003e emission.\u003c/p\u003e\n\u003cp\u003eIn substrate addition experiment, two sets of prepared jars as in warming experiment were incubated in aerobic and anaerobic conditions for 154 days to exhaust most labile carbon. Then three substrates with special \u003csup\u003e13\u003c/sup\u003eC were added into soil in powder:\u0026nbsp;oxalate\u0026nbsp;with \u0026delta;\u003csup\u003e13\u003c/sup\u003eC of -0.78 (AR,\u0026nbsp;\u0026ge;\u0026nbsp;99.5%, Macklin), glucose\u0026nbsp;with \u0026delta;\u003csup\u003e13\u003c/sup\u003eC of -12.43 (labeled glucose (U-\u003csup\u003e13\u003c/sup\u003eC, 99%, CIL) and glucose (AR\u0026nbsp;\u0026ge;\u0026nbsp;99.5%, Macklin) mixed by weight ratio of 1:1 ),\u0026nbsp;and\u0026nbsp;cinnamic acid\u0026nbsp;with \u0026delta;\u003csup\u003e13\u003c/sup\u003eC of -5.25 (AR,\u0026nbsp;\u0026ge;\u0026nbsp;99.5%, Macklin). The \u0026delta;\u003csup\u003e13\u003c/sup\u003eC of each substrate was determined before being added into soil (3 substrates * 3 depth * 2 conditions *3 replications = 54 jars). The\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e concentration of jars headspace was measured on the days of 1, 3, 5, 7, 12 and 17 after substrate addition. According to the CO\u003csub\u003e2\u003c/sub\u003e emission rate, the stable isotope \u003csup\u003e13\u003c/sup\u003eC\u0026nbsp;of respired\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e was measured on Day 12 after substrate addition when microbial growth was stable. The CO\u003csub\u003e2\u003c/sub\u003e emission rate and the CO\u003csub\u003e2\u003c/sub\u003e stable isotope \u003csup\u003e13\u003c/sup\u003eC\u0026nbsp;were also\u0026nbsp;measured\u0026nbsp;before substrate addition with a GasBench II system coupled to a Delta V Advantage IRMS (Thermo Scientific). Soil organic carbon (SOC) derived CO\u003csub\u003e2\u003c/sub\u003e (Priming effect) was calculated using equations1 and 2:\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/83400_b9e2661d18ef2d4b/83400_custom_files/img1631111302.png\"\u003e\u003c/p\u003e\n\u003cp\u003ewhere \u0026delta;\u003csup\u003e13\u003c/sup\u003eC\u003csub\u003eR\u003c/sub\u003e is the \u0026delta;\u003csup\u003e13\u003c/sup\u003eC of CO\u003csub\u003e2\u003c/sub\u003e from soil emission, \u0026delta;\u003csup\u003e13\u003c/sup\u003eC\u003csub\u003eso\u003c/sub\u003e\u003csub\u003ec\u003c/sub\u003e and \u0026delta;\u003csup\u003e13\u003c/sup\u003eC\u003csub\u003esub\u0026nbsp;\u003c/sub\u003eare the\u0026nbsp;\u0026delta;\u003csup\u003e13\u003c/sup\u003eC of soil and added substrates, and \u003cem\u003ef\u003c/em\u003e\u003csub\u003esom\u003c/sub\u003e and \u003cem\u003ef\u003c/em\u003e\u003csub\u003esub\u003c/sub\u003e are the contribution of soil and added substrates to the total amount of released CO\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e56\u003c/sup\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eS\u003c/strong\u003e\u003cstrong\u003eoil characteristics\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003ea\u003c/strong\u003e\u003cstrong\u003enalysis\u003c/strong\u003e\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eThe concentration of Fe\u0026nbsp;bound\u0026nbsp;organic carbon\u0026nbsp;(Fe-OC)\u0026nbsp;was quantitatively measured\u0026nbsp;on the basis of the citrate-bicarbonate-dithionite (CBD) method\u003csup\u003e57\u003c/sup\u003e\u003cem\u003e.\u003c/em\u003e Briefly, in a reduction treatment, a solution (15 ml, pH=7) containing 0.11 M sodium bicarbonate and 0.27 M trisodium citrate was added into 0.25 g freeze-dried soil and vortex oscillation in an 80\u0026deg;C environment heated by water bathing for 15 min. A reducing agent, sodium dithionite, was then added into mixture and 80\u0026deg;C water bathing for another 15 min. In the control treatment, instead of CBD extraction, soil was extracted with sodium chloride (NaCl) at an equivalent ionic strength. After centrifuging and filtering, the residual particles of both reduction and control treatments were rinsed with 5 ml DI water four times, freeze-dried, grounded to \u0026lt;100 \u0026mu;m and HCl-fumigated to remove the inorganic carbon, then were measured to get the SOC content. Fe-bound SOC content was calculated with the following equation:\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/83400_b9e2661d18ef2d4b/83400_custom_files/img1631111376.png\"\u003e\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"https://myfiles.space/user_files/83400_b9e2661d18ef2d4b/83400_custom_files/img1631111427.png\"\u003e\u003c/p\u003e\n\u003cp\u003eThe concentration of water-soluble phenolics in soil was estimated using the Folin-Ciocalteu method with \u003cem\u003ep\u003c/em\u003e-hydroxybenzoic acid as standard. Soil (5 g) and deionized water (25 ml) were shaken at a frequency of 200 rpm for 20 hours. After\u0026nbsp;filtering\u0026nbsp;through a 0.45\u0026nbsp;\u0026mu;m filter\u0026nbsp;and\u0026nbsp;diluting\u0026nbsp;1:5, the extracted solution (1 ml) was mixed with\u0026nbsp;2 ml\u0026nbsp;Na\u003csub\u003e2\u003c/sub\u003eCO\u003csub\u003e3\u003c/sub\u003e-NaHCO\u003csub\u003e3\u003c/sub\u003e buffer (pH 10), 1 ml Folin-Ciocalteu reagent, and 1 ml 0.4 mol l\u003csup\u003e-1\u003c/sup\u003e Na\u003csub\u003e2\u003c/sub\u003eCO\u003csub\u003e3\u003c/sub\u003e,and kept in\u0026nbsp;dark at 37\u0026deg;C for 15 min. Finally, the absorbance was measured at a wavelength of 750 nm using a spectrophotometer (Thermo Spectronic 200, USA). Standard phenolic absorbance was also measured\u0026nbsp;with the\u0026nbsp;same procedure. The concentration of water-soluble phenolics (measured in mg g\u003csup\u003e-1\u003c/sup\u003e soil) was calculated using absorbance values of the standard and the soil samples\u003csup\u003e58\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMicrob\u003c/strong\u003e\u003cstrong\u003eial analysis.\u0026nbsp;\u003c/strong\u003eMicrobial biomass carbon (MBC) concentration was measured using chloroform fumigation technique\u003csup\u003e59\u003c/sup\u003e. DNA\u0026nbsp;of three layers soil\u0026nbsp;was\u0026nbsp;extracted using a MoBio PowerSoil Kit (MoBio Laboratories, Carlsbad, CA, USA) following the manufacturer\u0026rsquo;s instructions. The V4 region of\u0026nbsp;bacterial 16S rRNA gene and the ITS1 region of fungi\u0026nbsp;gene\u0026nbsp;were amplified\u0026nbsp;during\u0026nbsp;PCR to characterize bacterial and fungal communities, respectively\u003csup\u003e60,61\u003c/sup\u003e. Quantitative PCR (qPCR) was performed to estimate the abundance of bacteria and fungi\u0026nbsp;of soil\u003csup\u003e62\u003c/sup\u003e.\u0026nbsp;The sequence data of 16S and ITS were retrieved from another paper by our team (deposited in the NCBI Sequence Read Archive (SRA) database under accession number\u0026nbsp;SRP211776).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEnzyme activity\u003c/strong\u003e\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eTwo oxidase\u0026nbsp;enzymes\u0026nbsp;(phenol oxidase\u0026nbsp;and\u0026nbsp;peroxidase)\u0026nbsp;were\u0026nbsp;assayed using the substrate\u0026nbsp;of\u0026nbsp;L-DOPA\u0026nbsp;and two hydrolase\u0026nbsp;were measured using the respective substrates\u0026nbsp;of\u0026nbsp;4-MUB-\u0026beta;-D-glucopyranoside and 4-MUB-\u0026beta;-D-cellobioside. Soil (1.25 g) was added to 50 mM acetate buffer (125 ml, pH 5.0) and shaken for 5 min at 200 rpm to make a suspension\u003csup\u003e63,64\u003c/sup\u003e for following analysis.\u003c/p\u003e\n\u003cp\u003eTwo hydrolase of\u0026nbsp;\u0026beta;-1,4-glucosidase and cellobiohydrolase were detected fluorometrically in the microplates. After filtering, 200 \u0026mu;l of sample solution was added to sample control, sample assay and quench tubes, while 200 \u0026mu;l of acetate buffer was added to blank, reference, and negative control tubes. Acetate buffer (50 \u0026mu;l) was also added to blank and sample controls, while 50 \u0026mu;l of 10 \u0026mu;M 4-methylumbelliferone substrate was added to reference and quench tubes, or 50 \u0026mu;l of 4-MUB-conjugated substrates\u0026nbsp;added\u0026nbsp;to negative control and sample assay tubes. All microplates were incubated at 20\u0026deg;C in the dark for four hours before the addition of 10 \u0026mu;l of 1.0 M NaOH to stop the reaction. Fluorescence was measured using a microplate fluorometer with 365 nm excitation and 450 nm emission filters. The fluorescence of the negative controls and the quenching were measured together and expressed in nmol h\u003csup\u003e-1\u003c/sup\u003e g\u003csup\u003e-1\u003c/sup\u003e.\u003c/p\u003e\n\u003cp\u003eTwo oxygen enzyme activities\u0026nbsp;were measured spectrophotometrically. Similar to the fluorometric analysis, 200 \u0026mu;l of filtered\u0026nbsp;soil\u0026nbsp;solution was added to\u0026nbsp;tubes of\u0026nbsp;sample control and sample assay; 200 \u0026mu;l of acetate buffer\u0026nbsp;was added\u0026nbsp;to\u0026nbsp;tubes of\u0026nbsp;blank and negative controls; 50 \u0026mu;l of acetate buffer\u0026nbsp;was add to\u0026nbsp;tubes of\u0026nbsp;blank and sample controls; and 50 \u0026mu;l of 25 mmol L-DOPA\u0026nbsp;was added\u0026nbsp;to\u0026nbsp;tubes of\u0026nbsp;negative control and sample assay. For peroxidase assays, another 10 \u0026mu;l of 0.3% H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e was added to each tube. All microplates were incubated at 20\u0026deg;C in dark for 20 hours. The absorbance was measured at 450 nm using a microplate spectrophotometer (Varioskan Flash, Thermo, USA). The activity was expressed in \u0026mu;mol h\u003csup\u003e-1\u003c/sup\u003e g\u003csup\u003e-1\u003c/sup\u003e.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData analysis\u003c/strong\u003e\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eDifferences in soil biogeochemical characteristics and soil\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission across soil layers were assessed for significance using ANOVA with Duncan test. ANOVA was also used to assess the differences of soil CO\u003csub\u003e2\u003c/sub\u003e emission when three substrates were added. \u003cem\u003et\u003c/em\u003e\u003cem\u003e-\u003c/em\u003etest was used to assess the differences in soil\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission between 8\u0026deg;C and 18\u0026deg;C. All statistical analyses were conducted in SPSS 20.0 for Windows (SPSS, Chicago, IL, USA) and all figures were created in Origin 8.0. Microbial raw sequence data was processed using QIIME2 (version 2018.11) \u003csup\u003e65\u003c/sup\u003eand diversity was performed using\u0026nbsp;R (Version 3.3.1).\u003c/p\u003e\n\u003cp\u003ePiecewise structure equation model\u0026nbsp;(SEM) was used to compare the direct and indirect effects of temperature, inputting substrate, soil microbes (MBC), enzymes, soil phenolics /nitrogen nutrient on soil CO\u003csub\u003e2\u003c/sub\u003e emission of each layer, using piecesiseSEMpackage in R. One specific hypothesized interactions included in the model was outlined in Supplementary Fig. 12. The temperature and inputting fresh substrate were specified as exogenous variables; soil enzymes, MBC, soil carbon components, and nitrogen nutrient were endogenous variables. Paths were included from temperature and inputting fresh substrate to all endogenous variables, then to soil CO\u003csub\u003e2\u003c/sub\u003e emission. The model was used in linear mixed effects models with block as a random effect in the piecewise SEM. A stepwise removal of the least significant paths from the models was conducted until either the AIC did not decrease with removal of the next parameter or there were no remaining parameters with \u003cem\u003ep\u003c/em\u003e \u0026gt; 0.1.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003eSoil properties.\u003c/strong\u003e Of the soil carbon quality proxies, the concentration of Fe-OC was\u0026nbsp;significantly higher at ETzone than at aerobic surface and anaerobic deep soil (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01),\u0026nbsp;reflecting the abundance of recalcitrant and aggregated protected carbon in ETzone. H\u003csub\u003e2\u003c/sub\u003eO-PhO was higher at ETzone and anaerobic deep soil than at aerobic surface soil (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01; Table 1). In contrast with soil carbon quality proxies, the microbial activities including soil MBC (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01), the Shannon diversity of bacteria (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01) and fungi (\u003cem\u003ep\u003c/em\u003e \u0026gt; 0.05), were significantly lower at ETzone than at aerobic surface and anaerobic deep soil. The qPCR results showed that bacterial\u0026nbsp;(\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01) and fungal (\u003cem\u003ep\u003c/em\u003e \u0026gt; 0.05) copy numbers were lower in anaerobic deep soil than in\u0026nbsp;aerobic surface soil. In terms of soil enzymes,\u0026nbsp;phenolic oxidase\u0026nbsp;(\u003cem\u003ep\u003c/em\u003e \u0026gt; 0.05) and\u0026nbsp;\u0026beta;-1-4-glucosidase\u0026nbsp;(\u003cem\u003ep\u003c/em\u003e \u0026gt; 0.05) had the highest activities at aerobic surface soil, while the highest activity of peroxidase (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05) and cellobiohydrolase\u0026nbsp;(\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01) were at ETzone (Table 1).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResponse\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;to warming\u003c/strong\u003e\u003cstrong\u003e.\u003c/strong\u003e After a long-term incubation, the potential CO\u003csub\u003e2\u003c/sub\u003e emission rate at ETzone\u0026nbsp;(28.31 \u0026plusmn; 3.55\u0026nbsp;\u0026mu;g\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e)\u0026nbsp;was significantly lower than the other two layers\u0026nbsp;(surface:\u0026nbsp;48.44 \u0026plusmn; 3.59\u0026nbsp;\u0026mu;g\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e; subsurface:\u0026nbsp;38.06 \u0026plusmn; 1.71\u0026nbsp;\u0026mu;g\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e;\u0026nbsp;\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01). With the lowest Q\u003csub\u003e10\u003c/sub\u003e (1.37) among the three layers, ETzone soil CO\u003csub\u003e2\u003c/sub\u003e emission also did not show significant increase (\u003cem\u003ep\u003c/em\u003e \u0026gt; 0.05) in warming treatment. Different from the ETzone, the CO\u003csub\u003e2\u003c/sub\u003e emission in aerobic surface and anaerobic deep soil were significantly enhanced\u0026nbsp;by warming (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01\u0026nbsp;for both; Fig 2a), with Q\u003csub\u003e10\u003c/sub\u003e of 2.14 and 1.51, respectively (Table 1). These results indicated that the soil at ETzone was not sensitive to warming. Warming also had significantly positive effect on MBC concentration of all three layers soil (Supplementary Fig.\u0026nbsp;4). With soil degrading from field anaerobic conditions to aerobic scenario, the warming response of CO\u003csub\u003e2\u003c/sub\u003e emission of both ETzone and anaerobic deep soil were significantly enhanced (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01 for both;\u0026nbsp;Fig 2a), with increments of\u0026nbsp;65.7% to 83.4%\u0026nbsp;at ETzone\u0026nbsp;(Supplementary Fig.\u0026nbsp;5a)\u0026nbsp;and 68.4% to 89.2% at anaerobic deep soil (Supplementary Fig.\u0026nbsp;5b). In future aerobic scenario, the activity of soil oxidase enzymes including\u0026nbsp;phenol oxidase and peroxidase were significantly higher than in field conditions and further increased by warming (Supplementary Fig. 6). From field anaerobic conditions to future degraded scenario, the increment of CO\u003csub\u003e2\u003c/sub\u003e emission facilitated by warming of the upper 100 cm soil profile increased by 33.7% (ranging\u0026nbsp;from 84.62\u0026nbsp;\u0026plusmn;\u0026nbsp;3.67\u0026nbsp;\u0026mu;g\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e to 127.92\u0026nbsp;\u0026plusmn;\u0026nbsp;7.38\u0026nbsp;\u0026mu;g\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e;\u0026nbsp;Supplementary Fig. 7).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResponse to p\u003c/strong\u003e\u003cstrong\u003eriming\u003c/strong\u003e\u003cstrong\u003e.\u003c/strong\u003e The response to priming of soil carbon emission varied with soil conditions and \u0026nbsp;layers.\u0026nbsp;In field conditions, new substrate addition increased soil CO\u003csub\u003e2\u003c/sub\u003e emission in all three layers, but significantly only at ETzone (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01), not at aerobic surface or anaerobic deep soil (\u003cem\u003ep\u003c/em\u003e \u0026gt; 0.05 for both), demonstrating that the ETzone was more sensitive to new substrate than the other two layers. In future degraded aerobic scenario,\u0026nbsp;the\u0026nbsp;increments of CO\u003csub\u003e2\u003c/sub\u003e emission caused by substrate addition were significant at\u0026nbsp;both\u0026nbsp;ETzone and anaerobic deep soil (\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01 for both; Fig\u0026nbsp;2b). Substrates like glucose and cinnamic acid also enriched fungal abundance (Supplementary Fig.\u0026nbsp;8), increased phenol oxidase and peroxidase activities significantly in both field conditions\u0026nbsp;and future degraded scenario (Supplementary Fig.\u0026nbsp;9).\u003c/p\u003e\n\u003cp\u003eThe priming response of soil carbon at each layer varied with substrates and conditions. In the field conditions, CO\u003csub\u003e2\u003c/sub\u003e emission was significantly enhanced by oxalate in the aerobic surface soil, by cinnamic acid in the anaerobic deep soil, and by both in the ETzone. Glucose did not show any significant influence on soil CO\u003csub\u003e2\u003c/sub\u003e emission in any layers (Fig 3a). In the degraded aerobic scenario, however, every substrate increased soil CO\u003csub\u003e2\u003c/sub\u003e emission of all three layers (Fig 3b).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eComparison of\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;priming\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;and warming\u003c/strong\u003e\u003cstrong\u003e.\u003c/strong\u003e The log values\u0026nbsp;of the increased CO\u003csub\u003e2\u003c/sub\u003e emission caused by priming and warming\u0026nbsp;were compared in\u0026nbsp;three layers, to predict the main factor influencing soil CO\u003csub\u003e2\u003c/sub\u003e emission.\u0026nbsp;In field conditions, we found that warming had a higher log value than priming at aerobic surface and anaerobic deep soil. However, at ETzone, the log value of priming was higher than warming. These\u0026nbsp;results indicated\u0026nbsp;that\u0026nbsp;aerobic surface and anaerobic deep soil\u0026nbsp;were more influenced by warming and the\u0026nbsp;ETzone\u0026nbsp;was more influenced by priming. In the future degraded scenario, the log value of\u0026nbsp;both\u0026nbsp;warming and priming increased, with different increments\u0026nbsp;at ETzone and anaerobic deep soil\u0026nbsp;(Fig 4).\u0026nbsp;Similar\u0026nbsp;increment\u0026nbsp;of warming and priming\u0026nbsp;at\u0026nbsp;anaerobic\u0026nbsp;deep\u0026nbsp;lead to the main influencing factor still was warming. At ETzone, greater increment of warming resulted in the main influecning factor became wamring\u0026nbsp;(Fig\u0026nbsp;4).\u0026nbsp;In terms of different substrates, each showed different priming log value for three layers, substrate with\u0026nbsp;strong priming effect\u0026nbsp;(Fig 3) had\u0026nbsp;higher\u0026nbsp;log value (Supplementary Fig.\u0026nbsp;10).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeterminants\u0026nbsp;of CO\u003csub\u003e2\u003c/sub\u003e emission.\u0026nbsp;\u003c/strong\u003eSoil variables varied significantly in warming and priming responses. The significantly correlated soil variables were differed among three layers (Supplementary Fig. 11).\u0026nbsp;SEM\u0026nbsp;results showed that soil variables could explain 75%, 75% and 61.1% of the variability of CO\u003csub\u003e2\u003c/sub\u003e emission of aerobic surface, ETzone and anaerobic deep soil, respectively (Fig 5). At aerobic surface soil, NH+ 4 (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01), MBC (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.05) and\u0026nbsp;\u0026beta;-1-4-glucosidase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05)\u0026nbsp;had positive effect on\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission. NH+ 4 also indirectly affect CO\u003csub\u003e2\u003c/sub\u003e emission through being negatively correlated with MBC\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05) and positively correlated with\u0026nbsp;\u0026beta;-1-4-glucosidase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01). Temperature and inputting substrate exerted strong indirect effect on\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission through their positive correlation with NH+ 4 (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.001) and MBC (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01;\u0026nbsp;Fig 5a).\u0026nbsp;For anaerobic deep soil,\u0026nbsp;cellobiohydrolase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01)\u0026nbsp;and MBC\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05)\u0026nbsp;had directly positive effect on\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission,\u0026nbsp;and H\u003csub\u003e2\u003c/sub\u003eO-PhO\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05)\u0026nbsp;had negative effect on\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission. Temperature exerted strong indirect effects on\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission through its positive correlation with MBC (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01), and the inputting substrate exerted indirect effect on CO\u003csub\u003e2\u003c/sub\u003e emission through its positive correlation with\u0026nbsp;cellobiohydrolase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.05)\u0026nbsp;and negative correlation with MBC\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01)\u0026nbsp;and\u0026nbsp;H\u003csub\u003e2\u003c/sub\u003eO-PhO\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.05;\u0026nbsp;Fig 5c). At ETzone, CO\u003csub\u003e2\u003c/sub\u003e emission was positively affected by soil\u0026nbsp;phenolic oxidase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01) and MBC (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05), but negatively by\u0026nbsp;H\u003csub\u003e2\u003c/sub\u003eO-PhO\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05). MBC also indirectly affected CO\u003csub\u003e2\u003c/sub\u003e emission through being negatively correlated with\u0026nbsp;H\u003csub\u003e2\u003c/sub\u003eO-PhO\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026gt; 0.05). Temperature and inputting substrate exerted strong indirect effect on\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission through the positive correlation with MBC (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.001),\u0026nbsp;phenolic oxidase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01)\u0026nbsp;and H\u003csub\u003e2\u003c/sub\u003eO-PhO (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01;\u0026nbsp;Fig 5b).\u0026nbsp;Taken together, the most important drivers of CO\u003csub\u003e2\u003c/sub\u003e emission were temperature (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.001) and NH+\u0026nbsp;4(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01) at aerobic surface soil, and were temperature, the inputting substrate and\u0026nbsp;cellobiohydrolase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01 for all) at anaerobic deep soil. The main drivers of CO\u003csub\u003e2\u003c/sub\u003e emission at ETzone were inputting substrate (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.001) and\u0026nbsp;phenolic oxidase\u0026nbsp;(\u003cem\u003ep\u0026nbsp;\u003c/em\u003e\u0026lt; 0.01; Fig 5).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe CO\u003csub\u003e2\u003c/sub\u003e emission at ETzone was significantly lower than the upper aerobic surface soil (41.6%) and anaerobic deep soil (34.4%). Moreover, 10\u0026deg;C warming failed to stimulate ETzone CO\u003csub\u003e2\u003c/sub\u003e emission significantly, while significantly stimulated that in the upper aerobic surface and anaerobic deep soil in the conditions of pristine field. In contrast with most other findings labeling ETzone as a hotspot in the ecosystem\u003csup\u003e1,4,5,34\u003c/sup\u003e, the lowest\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission rate and least warming response of the ETzone at the subsurface of peatland in our study showed that it was inert and resistant to warming. Such phenomenon can be explained by the constitution and characteristics of the carbon at ETzone soil. It is known that the ETzone in peatland experienced most severe disturbance during long-term water table fluctuation, which consumes great amounts of labile carbon and enriches abundance of Fe-OC (Table 1). Fe-OC is recalcitrant to soil microbes and warming resistant\u003csup\u003e36\u003c/sup\u003e.\u0026nbsp;Higher\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission from the aerobic surface and the anaerobic deep soil, consistent with other studies\u003csup\u003e13,14\u003c/sup\u003e, reflects\u0026nbsp;the presence of adequate\u0026nbsp;plant substrates\u0026nbsp;accessible to microbes\u003csup\u003e35\u003c/sup\u003e and high microbial activity\u003csup\u003e36\u003c/sup\u003e (Table 1, Supplementary Fig. 4). The relatively lower Fe-OC likely contributed to the higher sensitivity of the anaerobic deep soil to warming\u003csup\u003e37\u003c/sup\u003e (Table 1). The inert feature and non-responsive characteristic make ETzone a buffer against soil carbon loss from peatlands under warming climate, rather than a hotspot.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eHowever, the warming resistant characteristics of the ETzone was compromised when exposed to aerobic conditions simulating future degraded scenario. Our results showed that the CO\u003csub\u003e2\u003c/sub\u003e emission from the ETzone became significantly sensitive to warming, and the deeper adjacent soil became even more sensitive to warming under aerobic conditions. These results were consistent with a previous study indicating that drier or aerobic subsurface soil was sensitive to warming\u003csup\u003e10\u003c/sup\u003e. In our\u0026nbsp;study, the presence of O\u003csub\u003e2\u003c/sub\u003e increased the activity of the oxidase enzyme (Supplementary Fig. 6), which enhanced soil carbon decomposition\u003csup\u003e38\u003c/sup\u003e.\u0026nbsp;Compared to that of pristine field conditions, the increment of CO\u003csub\u003e2\u003c/sub\u003e emission caused by warming of the upper 100 cm soil profile was 33.7% higher in the degraded scenario. Therefore, about one third more CO\u003csub\u003e2\u0026nbsp;\u003c/sub\u003ewill be stabilized by the protection of the inert and warming-resistant ETzone, given it being kept in an anaerobic dominated condition (Supplementary Fig. 7).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eNoteworthy, contrary to the warming response, CO\u003csub\u003e2\u0026nbsp;\u003c/sub\u003eemission at\u0026nbsp;the ETzone was significantly sensitive to priming of fresh substrates,\u0026nbsp;even\u0026nbsp;under\u0026nbsp;anaerobic\u0026nbsp;conditions,\u0026nbsp;no matter labile carbon of oxalate and glucose, or recalcitrant carbon of cinnamic acid.\u0026nbsp;There are some possible reasons for this phenomenon.\u0026nbsp;Due to frequent water table fluctuation, SOC at ETzone was dominated in\u0026nbsp;mineral associations (Table 1) and the microbes were limited by\u0026nbsp;energy\u003csup\u003e33\u003c/sup\u003e.\u0026nbsp;Exo-substrates representing both labile and recalcitrant carbon alleviated\u0026nbsp;energy limitation\u0026nbsp;of the microbes, especially for\u0026nbsp;fungi\u003csup\u003e33\u003c/sup\u003e which were of relatively low abundance in original soils (Table 1) and increased greatly in diversity and relative abundance after substrate addition (Supplementary Fig. 8). Fungi could excrete substantial extracellular enzymes (Supplementary Fig. 9a and c) to decompose recalcitrant carbon and break organo-mineral complexes, liberating organic compounds from protected associations or aggregates\u003csup\u003e39,40\u003c/sup\u003e for further decomposition.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eDifferent from that of ETzone,\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission\u0026nbsp;from the aerobic surface soil remained\u0026nbsp;stable\u0026nbsp;after\u0026nbsp;substrate addition, except with oxalic acid (Fig\u0026nbsp;3). With\u0026nbsp;strong metal-complexing ability, oxalic acid can cause strong priming effect by\u0026nbsp;breaking\u0026nbsp;mineral-organic associations, but\u0026nbsp;is of limited bioenergetics use to microbes\u003csup\u003e39\u003c/sup\u003e.\u0026nbsp;The significant influence of oxalic acid\u0026nbsp;demonstrated that the surface soil\u0026nbsp;was not energy-limited.\u0026nbsp;In the anaerobic\u0026nbsp;deep soil,\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission was\u0026nbsp;also\u0026nbsp;enhanced by the addition of substrates, although\u0026nbsp;not significantly (Fig\u0026nbsp;2b), except\u0026nbsp;for the addition of\u0026nbsp;cinnamic acid\u0026nbsp;(Fig\u0026nbsp;3), which contains an aromatic loop and is a complex\u0026nbsp;product of lignin depolymerization\u003csup\u003e32\u003c/sup\u003e. The significant increase in soil CO\u003csub\u003e2\u003c/sub\u003e emission caused by cinnamic acid suggested\u0026nbsp;that the deep soil was\u0026nbsp;rich\u0026nbsp;in recalcitrant carbon in the form of aggregates\u003csup\u003e36\u003c/sup\u003e.\u0026nbsp;The protection offered by aggregates rendered the soil insensitive to\u0026nbsp;other substrates.\u0026nbsp;Combined with\u0026nbsp;aerobic conditions\u0026nbsp;which opened\u0026nbsp;the enzyme \u0026ldquo;latch\u0026rdquo;\u003csup\u003e38\u003c/sup\u003e (Supplementary Fig. 9b and\u0026nbsp;d),\u0026nbsp;the\u0026nbsp;priming effects\u0026nbsp;of\u0026nbsp;all substrates\u0026nbsp;were enhanced\u0026nbsp;in the degraded\u0026nbsp;scenario, leading to further decomposition of liberated organic compounds\u0026nbsp;(Fig\u0026nbsp;3b).\u003c/p\u003e\n\u003cp\u003eWe found that the response of the ETzone soil to warming and priming significantly differed from the other two layers. Under\u0026nbsp;the\u0026nbsp;field conditions, the aerobic surface soil was influenced more\u0026nbsp;intensively\u0026nbsp;by warming than by\u0026nbsp;priming (Fig 4), which probably reflected that the activity of soil microbes and enzymes were suppressed by\u0026nbsp;low temperature\u003csup\u003e36\u003c/sup\u003e.\u0026nbsp;Warming could enhance soil carbon decomposition by favoring\u0026nbsp;soil microbes and enzyme activity\u003csup\u003e36,41\u003c/sup\u003e.\u0026nbsp;Similarly, the anaerobic deep soil\u0026nbsp;was\u0026nbsp;also\u0026nbsp;influenced\u0026nbsp;more intensively\u0026nbsp;by warming than\u0026nbsp;priming, though\u0026nbsp;with\u0026nbsp;a\u0026nbsp;weaker\u0026nbsp;response to warming than the surface soil\u0026nbsp;(Fig 4). This\u0026nbsp;is\u0026nbsp;ascribed\u0026nbsp;to the\u0026nbsp;long-term\u0026nbsp;anaerobic conditions\u0026nbsp;and\u0026nbsp;the\u0026nbsp;limited\u0026nbsp;soil nutrients in the deep soil\u003csup\u003e42\u003c/sup\u003e (Table 1). Even when soil conditions changed from the field anaerobic to the future degraded aerobic, the dominant effect at anaerobic deep soil kept as warming.\u003c/p\u003e\n\u003cp\u003eWith the strongest priming effect among the three layers,\u0026nbsp;the dominant\u0026nbsp;effect\u0026nbsp;for\u0026nbsp;ETzone was priming.\u0026nbsp;Soil carbon in the\u0026nbsp;ETzone\u0026nbsp;was\u0026nbsp;substantially\u0026nbsp;stabilized\u0026nbsp;due to\u0026nbsp;the\u0026nbsp;abundant\u0026nbsp;recalcitrant carbon inaccessible for microbes, e.g., Fe-OC\u003csup\u003e36\u003c/sup\u003e (Table 1), and the microbes were limited in nutruent. Substrate addition\u0026nbsp;helped\u0026nbsp;mitigate this limitation by supplying\u0026nbsp;carbon and\u0026nbsp;energy to soil microbes\u003csup\u003e43\u003c/sup\u003e. When\u0026nbsp;soil conditions\u0026nbsp;changed\u0026nbsp;from\u0026nbsp;the field\u0026nbsp;anaerobic to\u0026nbsp;the future degraded\u0026nbsp;aerobic,\u0026nbsp;however, the dominant effect for ETzone changed\u0026nbsp;from\u0026nbsp;priming\u0026nbsp;to warming\u0026nbsp;(Fig 4).\u0026nbsp;In oxygen unlimited environment, aerobic decomposition could releases nutrient from soil aggregates and recalcitrant carbon, easing the limitation of energy and nutrient on soil microbes. Thus, even with increase of effect by both warming and priming, priming was no longer the dominant factor.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main determinants of soil CO\u003csub\u003e2\u003c/sub\u003e emission at ETzone also differed with the other two layers. Our result showed that aerobic surface soil CO\u003csub\u003e2\u003c/sub\u003e emission was significantly determined by NH+ 4, demonstrating that surface soil was in N-limited conditions, consisting with previous researches which showed that peatlands on Zoige was a nitrogen limited system\u003csup\u003e44\u003c/sup\u003e. As with observed in warming response, temperature was indeed one of critical factors in determining aerobic surface soil CO\u003csub\u003e2\u003c/sub\u003e emission, through indirectly influencing soil microbial biomass\u003csup\u003e45\u003c/sup\u003e (Fig 5). For CO\u003csub\u003e2\u003c/sub\u003e emission from anaerobic deep soil, temperature was a critical factor too\u003csup\u003e15\u003c/sup\u003e, since warming could increase soil microbial biomass and\u0026nbsp;cellobiohydrolase, which are limited by temperature at both surface and deep soil\u003csup\u003e45,46\u003c/sup\u003e. Soil microbes at these two layers might also be in energy limited conditions, for their activities increased significantly after fresh substrate addition\u003csup\u003e33\u003c/sup\u003e.\u0026nbsp;The significant direct influence of cellobiohydrolase\u0026nbsp;at anaerobic deep soil demonstrated that the carbon in this layer was more complex than in aerobic surface soil\u003csup\u003e36\u003c/sup\u003e.\u0026nbsp;Contrasted with that at aerobic surface and anaerobic deep soil, the CO\u003csub\u003e2\u003c/sub\u003e emission at ETzone was mainly determined by\u0026nbsp;phenolic oxidase\u0026nbsp;(Fig 5), indicating that substantial recalcitrant carbon subject to phenolic oxidase existed at ETzone, and that the released CO\u003csub\u003e2\u003c/sub\u003e may mainly result from the decomposition of these recalcitrant carbon. Phenolic oxidase\u0026nbsp;and microbes at ETzone were limited in energy and nutrient, and stimulated by fresh carbon. The limitation of soil phenolic oxidase\u0026nbsp;and microbe at ETzone could better explain the significant response of ETzone CO\u003csub\u003e2\u003c/sub\u003e emission to added substrates.\u003c/p\u003e\n\u003cp\u003eFrom the\u0026nbsp;CO\u003csub\u003e2\u003c/sub\u003e emission\u0026nbsp;characteristics and its major determinants, we can see that the ETzone of the subsurface soil was characterized with both\u0026nbsp;warming-resistance and\u0026nbsp;substrate-sensitivity\u0026nbsp;(Fig 6). The warming-resistant ETzone\u0026nbsp;could serve as a buffer to\u0026nbsp;prevent depletion of carbon\u0026nbsp;stock inside and below peat due to decomposition in a warming\u0026nbsp;climate. For example, for northern peatlands with a carbon stock of 547 Pg (an average depth of 230 cm)\u003csup\u003e47,48\u003c/sup\u003e, and an ETzone presumably of 23.3 cm below the peatland surface (Fig 1), the inert ETzone would protect about 89.9% of the carbon stored inside and below it from warming-induced decomposition. However, due to the priming-sensitiveness of the ETzone, such protection\u0026nbsp;can easily\u0026nbsp;collapse through priming.\u0026nbsp;Thus, a non-priming ETzone will be beneficial to maintain carbon stability of peatlands. A\u0026nbsp;community with\u0026nbsp;stable\u0026nbsp;plant\u0026nbsp;composition\u0026nbsp;produces relatively constant amounts\u0026nbsp;and type of root\u0026nbsp;exudates\u003csup\u003e49,50\u003c/sup\u003e, and\u0026nbsp;stable\u0026nbsp;hydrological conditions\u0026nbsp;are helpful for\u0026nbsp;minimizing substrate exchange among soil layers\u003csup\u003e51\u003c/sup\u003e and the\u0026nbsp;positive\u0026nbsp;priming effects on\u0026nbsp;soil carbon,\u0026nbsp;ultimately decreases carbon loss\u003csup\u003e52\u003c/sup\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eWith ever more severe disturbance of climate change and human activities, carbon loss from\u0026nbsp;peatlands\u0026nbsp;had substantially increased in\u0026nbsp;the\u0026nbsp;last few decades\u003csup\u003e53,54\u003c/sup\u003e.\u0026nbsp;Several mechanisms were proposed as the key to keep peatlands carbon stable, e.g., the enzyme\u0026nbsp;\u0026ldquo;latch\u0026rdquo;\u0026nbsp;theory that emphasizes the importance of\u0026nbsp;anaerobic environment\u003csup\u003e38\u003c/sup\u003e in limiting enzyme activities and suggests\u0026nbsp;sustaining a stable\u0026nbsp;and\u0026nbsp;high water table\u0026nbsp;to protect carbon stock; and the \u0026ldquo;iron\u0026nbsp;gate\u0026rdquo;\u0026nbsp;theory about the essential role of Fe-OC to decrease carbon decomposition and protection of soil aggregation\u003csup\u003e40,55\u003c/sup\u003e. In our study, we found that maintaining a relatively constant status of the ETzone at the subsurface soil is critical in protecting peatland carbon stock. Because of the priming-sensitiveness of ETzone, the priority is to\u0026nbsp;establish a non-priming\u0026nbsp;ETzone\u0026nbsp;by\u0026nbsp;maintaining intact vegetation composition to\u0026nbsp;reduce\u0026nbsp;the\u0026nbsp;possibility of intensified positive\u0026nbsp;priming effect;\u0026nbsp;Furthermore, due to alleviation in\u0026nbsp;warming-resistance\u0026nbsp;of the ETzone soil in the degraded scenario, another important step is to\u0026nbsp;sustain\u0026nbsp;the depth of warming resistant ETzone through maintaining a constant water regime (Fig 6).\u0026nbsp;Our results also illustrated that ETzones along environmental gradients will not always be \u0026ldquo;hotspot\u0026rdquo; of biogeochemical processes and interactions, but could be a \u0026ldquo;buffer\u0026rdquo; against carbon loss. However, whether this theory is applicable across all terrestrial ecosystems\u0026nbsp;needs further investigation. Considering the limitation of the incubation experiment that missed the information of seasonal variations in temperature, phenology and soil moisture, there was still much uncertainty in illuminating \u003cem\u003ein situ\u003c/em\u003e carbon dynamic.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eOur results provided a compelling evidence of the stabilizing function of the ETzone on peatland carbon stock. Previous studies that had highlighted the critical role of subsoil in peatlands carbon dynamic, without considering the buffer capacity of the ETzone, may have overestimated the carbon loss under warming climate of peatlands, and underestimated the carbon loss through priming by shift in vegetation composition under climate change and human disturbance. Moreover, including the function of the ETzone in ecological modeling will be beneficial to accurately simulate the impacts of global change on SOC dynamic of peatlands. With climate change and the induced deeper plant root distributing, the stabilization of the ETzone would be non-negligible in predicting peatlands carbon dynamics.\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe appreciate the assistance in language editing by Ms. Wan Xiong.\u0026nbsp;This work was supported by the Strategic Priority Research Program of Chinese Academy of Science (XDA200500404), the Second TIBETAN Plateau Scientific Expedition (2019QZKK0304) and Key Research Program of Frontier Sciences, CAS, QYZDB-SSW-DQC007.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003cstrong\u003e:\u003c/strong\u003e H.C,\u0026nbsp;L.L, N.W, H.W, and Y.W\u0026nbsp;designed\u0026nbsp;the experiments.\u0026nbsp;L.L, J.T, and D.X performed the experiment.\u0026nbsp;L.L and H.C\u0026nbsp;analyzed the data.\u0026nbsp;L.L,\u0026nbsp;H.C, Y.W, N.W, X.X, H.W, C. P, and M.W wrote the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAdditional information\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSupplementary Materials:\u0026nbsp;Supplementary Figures.\u0026nbsp;1\u0026nbsp;to 12\u003c/p\u003e\n\u003cp\u003eCorrespondence and requests for materials should be addressed to N.W and Y.W.\u003c/p\u003e"},{"header":"References","content":"\u003cp\u003e1 Juutinen, S.\u003cem\u003e\u0026nbsp;et al.\u003c/em\u003e Major implication of the littoral zone for methane release from boreal lakes. \u003cem\u003eGlobal Biogeochemical Cycles\u003c/em\u003e\u003cstrong\u003e17\u003c/strong\u003e, doi:10.1029/2003GB002105, doi:10.1029/2003gb002105 (2003).\u003c/p\u003e\n\u003cp\u003e2 Pegoraro, E.\u003cem\u003e\u0026nbsp;et al.\u003c/em\u003e Glucose addition increases the magnitude and decreases the age of soil respired carbon in a long-term permafrost incubation study. \u003cem\u003eSoil Biology and Biochemistry\u003c/em\u003e\u003cstrong\u003e129\u003c/strong\u003e, 201-211, doi:10.1016/j.soilbio.2018.10.009 (2019).\u003c/p\u003e\n\u003cp\u003e3 Riedel, T., Zak, D., Biester, H. \u0026amp; Dittmar, T. 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G.\u003cem\u003e\u0026nbsp;et al.\u003c/em\u003e QIIME allows analysis of highthroughput community sequencing data. \u003cem\u003eNature methods\u003c/em\u003e\u003cstrong\u003e7\u003c/strong\u003e, 335-336, doi:10.1038/nmeth0510-335 (2010).\u003c/p\u003e"},{"header":"Tables","content":"\u003cp\u003e\u003cstrong\u003eTable 1\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eSoil properties of original samples from the three soil\u0026nbsp;layers\u0026nbsp;on Zoige peatland.\u003c/p\u003e\n\u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003eAerobic\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003eETzone\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003eAnaerobic\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003e\u003cstrong\u003eSoil characteristics\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eFe-OC (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e3.65 \u0026plusmn; 0.11B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e5.36 \u0026plusmn; 0.68A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e2.75 \u0026plusmn; 0.06B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eH\u003csub\u003e2\u003c/sub\u003eO-PhO\u0026nbsp;(mg kg\u003csup\u003e-1\u003c/sup\u003e)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e191.9 \u0026plusmn; 6.39B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e357.76 \u0026plusmn; 13.199A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e382.26 \u0026plusmn; 34.37A\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003e\u003cstrong\u003eDecompos\u003c/strong\u003e\u003cstrong\u003eer\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e \u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eMBC (mg kg\u003csup\u003e-1\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e3119.1 \u0026plusmn; 256.78A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e289.89 \u0026plusmn; 33.6C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e1262.37 \u0026plusmn; 111.33B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eBacteria Shannon diversity [\u003cem\u003eH\u003c/em\u003e\u003cem\u003e\u003csup\u003e、\u003c/sup\u003e\u003c/em\u003e]\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e5.59 \u0026plusmn; 0.15A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e4.33 \u0026plusmn; 0.17C\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e5.07 \u0026plusmn; 0.11B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eBacteria numbers (x10\u003csup\u003e6\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e142.29 \u0026plusmn; 21.31A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e29.46 \u0026plusmn; 5.21B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e20.97 \u0026plusmn; 3.2B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eFungi Shannon diversity [\u003cem\u003eH\u003c/em\u003e\u003cem\u003e\u003csup\u003e、\u003c/sup\u003e\u003c/em\u003e]\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e2.22 \u0026plusmn; 0.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e1.95 \u0026plusmn; 0.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e2.09 \u0026plusmn; 0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eFungi numbers (x10\u003csup\u003e6\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e3.79 \u0026plusmn; 2.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e0.13 \u0026plusmn; 0.07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e0.17 \u0026plusmn; 0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003ePhenolic oxidase (\u0026mu;mol h\u003csup\u003e-1\u003c/sup\u003e g\u003csup\u003e-1\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e5.8 \u0026plusmn; 0.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e4.87 \u0026plusmn; 0.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e3.93 \u0026plusmn; 0.72\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003ePeroxidase (\u0026mu;mol h\u003csup\u003e-1\u003c/sup\u003e g\u003csup\u003e-1\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e3.08 \u0026plusmn; 0.41ab\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e3.8 \u0026plusmn; 0.07a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e2.51 \u0026plusmn; 0.08b\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003e\u0026beta;-1-4-glucosidase (nmol h\u003csup\u003e-1\u003c/sup\u003e g\u003csup\u003e-1\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e5.5 \u0026plusmn; 0.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e4.97 \u0026plusmn; 0.6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e3.61 \u0026plusmn; 0.59\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eCellobiohydrolase\u0026nbsp;(nmol h\u003csup\u003e-1\u003c/sup\u003e g\u003csup\u003e-1\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e57.81 \u0026plusmn; 6.15A\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e71.38 \u0026plusmn; 9.33A\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e9.27 \u0026plusmn; 2.24B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003e\u003cstrong\u003eCarbon emission\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eCO\u003csub\u003e2\u003c/sub\u003e rate (\u0026mu;g CO\u003csub\u003e2\u003c/sub\u003e g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e)\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e48.44 \u0026plusmn; 3.59A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e28.31 \u0026plusmn; 3.55B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e38.06 \u0026plusmn; 1.71A\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"35.202492211838006%\"\u003e\n \u003cp\u003eQ\u003csub\u003e10\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e2.14 \u0026plusmn; 0.13A\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"20.560747663551403%\"\u003e\n \u003cp\u003e1.37 \u0026plusmn; 0.13B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.118380062305295%\"\u003e\n \u003cp\u003e1.51 \u0026plusmn; 0.08B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eValues are mean\u0026nbsp;\u0026plusmn;\u0026nbsp;SE (n = 3).\u0026nbsp;Fe-OC: Fe-bound soil organic carbon;\u0026nbsp;H\u003csub\u003e2\u003c/sub\u003eO-PhO: Water-soluble phenolics; MBC: microbial biomass carbon. Letters indicated the significant difference of soil properties among three layers (Duncan test; \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05 for lowercase and \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01 for uppercase).\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"environment, transitional zone, buffers, peatland carbon stock, biogeochemical cycle, climate change, intact vegetation composition","lastPublishedDoi":"10.21203/rs.3.rs-849966/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-849966/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eAn environment transitional zone (ETzone) is usually deemed as a hotspot in biogeochemical cycle, but little is known about its response to climate change. A typical ETzone develops at the subsurface of peatland after experiencing long-term water table fluctuation, characterized by alternative aerobic and anaerobic conditions. By an extensive incubation, we found that the CO\u003csub\u003e2\u003c/sub\u003e emission at this ETzone was 28.31 ± 3.55 μg g\u003csup\u003e-1\u003c/sup\u003e d\u003csup\u003e-1\u003c/sup\u003e, 41.6% and 34.4% lower than the upper (aerobic) and lower (anaerobic) layers, respectively. Moreover, with a lowest Q\u003csub\u003e10\u003c/sub\u003e of 1.37, its CO\u003csub\u003e2\u003c/sub\u003e emission was also the least warming-responsive, which could reduce 33.7% CO\u003csub\u003e2\u003c/sub\u003e loss in warming scenario. This result clearly revealed that the ETzone worked as a buffer to retard carbon loss, rather than a hotspot. Surprisingly, this buffer capacity of ETzone was easily collapse if being primed by fresh carbon. Therefore, maintaining a relatively stable ETzone is critical for protecting peatland carbon stock, and the priority is to block priming effect through maintaining an intact vegetation composition.\u003c/p\u003e","manuscriptTitle":"An Environment Transitional Zone Buffers Peatlands Carbon Loss","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2021-09-08 21:41:38","doi":"10.21203/rs.3.rs-849966/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"f17a42f9-a41d-4b30-8a75-6591ab25ddf6","owner":[],"postedDate":"September 8th, 2021","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":7051327,"name":"Planetary Science"},{"id":7051328,"name":"Environmental Chemistry"},{"id":7051329,"name":"Environmental Policy"}],"tags":[],"updatedAt":"2022-04-05T08:26:04+00:00","versionOfRecord":[],"versionCreatedAt":"2021-09-08 21:41:38","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-849966","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-849966","identity":"rs-849966","version":["v1"]},"buildId":"_2-kVJe1T_tPrBINL-cwx","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}