Experimental warming altered plant functional traits and their coordination in a permafrost ecosystem.

Knowledge about changes in plant functional traits is valuable for the mechanistic understanding of warming effects on ecosystem functions. However, observations have tended to focus on aboveground plant traits, and there is little information about changes in belowground plant traits or the coordination of above- and belowground traits under climate warming, particularly in permafrost ecosystems. Based on a 7-yr field warming experiment, we measured 26 above- and belowground plant traits of four dominant species, and explored community functional composition and trait networks in response to experimental warming in a permafrost ecosystem on the Tibetan Plateau. Experimental warming shifted community-level functional traits toward more acquisitive values, with earlier green-up, greater plant height, larger leaves, higher photosynthetic resource-use efficiency, thinner roots, and greater specific root length and root nutrient concentrations. However, warming had a negligible effect in terms of functional diversity. In addition, warming shifted hub traits which have the highest centrality in the network from specific root area to leaf area. These results demonstrate that above- and belowground traits exhibit consistent adaptive strategies, with more acquisitive traits in warmer environments. Such changes could provide an adaptive advantage for plants in response to environmental change.

Microbial nitrogen and phosphorus co-limitation across permafrost region.

The status of plant and microbial nutrient limitation have profound impacts on ecosystem carbon cycle in permafrost areas, which store large amounts of carbon and experience pronounced climatic warming. Despite the long-term standing paradigm assumes that cold ecosystems primarily have nitrogen deficiency, large-scale empirical tests of microbial nutrient limitation are lacking. Here we assessed the potential microbial nutrient limitation across the Tibetan alpine permafrost region, using the combination of enzymatic and elemental stoichiometry, genes abundance and fertilization method. In contrast with the traditional view, the four independent approaches congruently detected widespread microbial nitrogen and phosphorus co-limitation in both the surface soil and deep permafrost deposits, with stronger limitation in the topsoil. Further analysis revealed that soil resources stoichiometry and microbial community composition were the two best predictors of the magnitude of microbial nutrient limitation. High ratio of available soil carbon to nutrient and low fungal/bacterial ratio corresponded to strong microbial nutrient limitation. These findings suggest that warming-induced enhancement in soil nutrient availability could stimulate microbial activity, and probably amplify soil carbon losses from permafrost areas.

Accelerated organic matter decomposition in thermokarst lakes upon carbon and phosphorus inputs.

Mineralization of dissolved organic matter (DOM) in thermokarst lakes plays a non-negligible role in the permafrost carbon (C) cycle, but remains poorly understood due to its complex interactions with external C and nutrient inputs (i.e., aquatic priming and nutrient effects). Based on large-scale lake sampling and laboratory incubations, in combination with 13 C-stable-isotope labeling, optical spectroscopy, and high-throughput sequencing, we examined large-scale patterns and dominant drivers of priming and nutrient effects of DOM biodegradation across 30 thermokarst lakes along a 1100-km transect on the Tibetan Plateau. We observed that labile C and phosphorus (P) rather than nitrogen (N) inputs stimulated DOM biodegradation, with the priming and P effects being 172% and 451% over unamended control, respectively. We also detected significant interactive effects of labile C and nutrient supply on DOM biodegradation, with the combined labile C and nutrient additions inducing stronger microbial mineralization than C or nutrient treatment alone, illustrating that microbial activity in alpine thermokarst lakes is co-limited by both C and nutrients. We further found that the aquatic priming was mainly driven by DOM quality, with the priming intensity increasing with DOM recalcitrance, reflecting the limitation of external C as energy sources for microbial activity. Greater priming intensity was also associated with higher community-level ribosomal RNA gene operon (rrn) copy number and bacterial diversity as well as increased background soluble reactive P concentration. In contrast, the P effect decreased with DOM recalcitrance as well as with background soluble reactive P and ammonium concentrations, revealing the declining importance of P availability in mediating DOM biodegradation with enhanced C limitation but reduced nutrient limitation. Overall, the stimulation of external C and P inputs on DOM biodegradation in thermokarst lakes would amplify C-climate feedback in this alpine permafrost region.

Priming effect stimulates carbon release from thawed permafrost.

Climate warming leads to widespread permafrost thaw with a fraction of the thawed permafrost carbon (C) being released as carbon dioxide (CO2 ), thus triggering a positive permafrost C-climate feedback. However, large uncertainty exists in the size of this model-projected feedback, partly owing to the limited understanding of permafrost CO2 release through the priming effect (i.e., the stimulation of soil organic matter decomposition by external C inputs) upon thaw. By combining permafrost sampling from 24 sites on the Tibetan Plateau and laboratory incubation, we detected an overall positive priming effect (an increase in soil C decomposition by up to 31%) upon permafrost thaw, which increased with permafrost C density (C storage per area). We then assessed the magnitude of thawed permafrost C under future climate scenarios by coupling increases in active layer thickness over half a century with spatial and vertical distributions of soil C density. The thawed C stocks in the top 3 m of soils from the present (2000-2015) to the future period (2061-2080) were estimated at 1.0 (95% confidence interval (CI): 0.8-1.2) and 1.3 (95% CI: 1.0-1.7) Pg (1 Pg = 1015 g) C under moderate and high Representative Concentration Pathway (RCP) scenarios 4.5 and 8.5, respectively. We further predicted permafrost priming effect potential (priming intensity under optimal conditions) based on the thawed C and the empirical relationship between the priming effect and permafrost C density. By the period 2061-2080, the regional priming potentials could be 8.8 (95% CI: 7.4-10.2) and 10.0 (95% CI: 8.3-11.6) Tg (1 Tg = 1012 g) C year-1 under the RCP 4.5 and RCP 8.5 scenarios, respectively. This large CO2 emission potential induced by the priming effect highlights the complex permafrost C dynamics upon thaw, potentially reinforcing permafrost C-climate feedback.

Stochastic processes regulate belowground community assembly in alpine grasslands on the Tibetan Plateau.

Understanding biogeographical patterns and underlying processes of belowground community assembly is crucial for predicting soil functions and their responses to global environmental change. However, little is known about potential differences of belowground community assembly among bacteria, fungi, protists and soil animals, particularly for alpine ecosystems. Based on the combination of large-scale field sampling, high-throughput marker-gene sequencing and multiple statistical analyses, we explored patterns and drivers of belowground community assembly in alpine grasslands on the Tibetan Plateau. Our results revealed that the distance-decay rates varied among trophic levels, with organisms of higher trophic level having weaker distance-decay pattern. The spatial and environmental variables explained limited variations of belowground communities. By contrast, the stochastic processes, mainly consisting of dispersal limitation and drift, played a primary role in regulating belowground community assembly. Moreover, the relative importance of stochastic processes varied among trophic levels, with the role of dispersal limitation weakening whereas that of drift enhancing in the order of bacteria, fungi, protists and soil animals. These findings advance our understanding of patterns and mechanisms driving belowground community assembly in alpine ecosystems and provide a reference basis for predicting the dynamics of ecosystem functions under changing environment.

Nitrogen input enhances microbial carbon use efficiency by altering plant-microbe-mineral interactions.

Microbial growth and respiration are at the core of the soil carbon (C) cycle, as these microbial physiological performances ultimately determine the fate of soil C. Microbial C use efficiency (CUE), a critical metric to characterize the partitioning of C between microbial growth and respiration, thus controls the sign and magnitude of soil C-climate feedback. Despite its importance, the response of CUE to nitrogen (N) input and the relevant regulatory mechanisms remain poorly understood, leading to large uncertainties in predicting soil C dynamics under continuous N input. By combining a multi-level field N addition experiment with a substrate-independent 18 O-H2 O labelling approach as well as high-throughput sequencing and mineral analysis, here we elucidated how N-induced changes in plant-microbial-mineral interactions drove the responses of microbial CUE to N input. We found that microbial CUE increased significantly as a consequence of enhanced microbial growth after 6-year N addition. In contrast to the prevailing view, the elevated microbial growth and CUE were not mainly driven by the reduced stoichiometric imbalance, but strongly associated with the increased soil C accessibility from weakened mineral protection. Such attenuated organo-mineral association was further linked to the N-induced changes in the plant community and the increased oxalic acid in the soil. These findings provide empirical evidence for the tight linkage between mineral-associated C dynamics and microbial physiology, highlighting the need to disentangle the complex plant-microbe-mineral interactions to improve soil C prediction under anthropogenic N input.

Divergent Trajectory of Soil Autotrophic and Heterotrophic Respiration upon Permafrost Thaw.

Warming-induced permafrost thaw may stimulate soil respiration (Rs) and thus cause a positive feedback to climate warming. However, due to the limited in situ observations, it remains unclear about how Rs and its autotrophic (Ra) and heterotrophic (Rh) components change upon permafrost thaw. Here we monitored variations in Rs and its components along a permafrost thaw sequence on the Tibetan Plateau, and explored the potential linkage of Rs components (i.e., Ra and Rh) with biotic (e.g., plant functional traits and soil microbial diversity) and abiotic factors (e.g., substrate quality). We found that Ra and Rh exhibited divergent responses to permafrost collapse: Ra increased with the time of thawing, while Rh exhibited a hump-shaped pattern along the thaw sequence. We also observed different drivers of thaw-induced changes in the ratios of Ra:Rs and Rh:Rs. Except for soil water status, plant community structure, diversity, and root properties explained the variation in Ra:Rs ratio, soil substrate quality and microbial diversity were key factors associated with the dynamics of Rh:Rs ratio. Overall, these findings demonstrate divergent patterns and drivers of Rs components as permafrost thaw prolongs, which call for considerations in Earth system models for better forecasting permafrost carbon-climate feedback.

Large-scale evidence for microbial response and associated carbon release after permafrost thaw.

Permafrost thaw could trigger the release of greenhouse gases through microbial decomposition of the large quantities of carbon (C) stored within frozen soils. However, accurate evaluation of soil C emissions from thawing permafrost is still a big challenge, partly due to our inadequate understanding about the response of microbial communities and their linkage with soil C release upon permafrost thaw. Based on a large-scale permafrost sampling across 24 sites on the Tibetan Plateau, we employed meta-genomic technologies (GeoChip and Illumina MiSeq sequencing) to explore the impacts of permafrost thaw (permafrost samples were incubated for 11 days at 5°C) on microbial taxonomic and functional communities, and then conducted a laboratory incubation to investigate the linkage of microbial taxonomic and functional diversity with soil C release after permafrost thaw. We found that bacterial and fungal α diversity decreased, but functional gene diversity and the normalized relative abundance of C degradation genes increased after permafrost thaw, reflecting the rapid microbial response to permafrost thaw. Moreover, both the microbial taxonomic and functional community structures differed between the thawed permafrost and formerly frozen soils. Furthermore, soil C release rate over five month incubation was associated with microbial functional diversity and C degradation gene abundances. By contrast, neither microbial taxonomic diversity nor community structure exhibited any significant effects on soil C release over the incubation period. These findings demonstrate that permafrost thaw could accelerate C emissions by altering the function potentials of microbial communities rather than taxonomic diversity, highlighting the crucial role of microbial functional genes in mediating the responses of permafrost C cycle to climate warming.

Phosphorus rather than nitrogen regulates ecosystem carbon dynamics after permafrost thaw.

Ecosystem carbon (C) dynamics after permafrost thaw depends on more than just climate change since soil nutrient status may also impact ecosystem C balance. It has been advocated that nitrogen (N) release upon permafrost thaw could promote plant growth and thus offset soil C loss. However, compared with the widely accepted C-N interactions, little is known about the potential role of soil phosphorus (P) availability. We combined 3-year field observations along a thaw sequence (constituted by four thaw stages, i.e., non-collapse and 5, 14, and 22 years since collapse) with an in-situ fertilization experiment (included N and P additions at the level of 10 g N m-2  year-1 and 10 g P m-2  year-1 ) to evaluate ecosystem C-nutrient interactions upon permafrost thaw. We found that changes in soil P availability rather than N availability played an important role in regulating gross primary productivity and net ecosystem productivity along the thaw sequence. The fertilization experiment confirmed that P addition had stronger effects on plant growth than N addition in this permafrost ecosystem. These two lines of evidence highlight the crucial role of soil P availability in altering the trajectory of permafrost C cycle under climate warming.

Above- and below-ground resource acquisition strategies determine plant species responses to nitrogen enrichment.

BACKGROUND AND AIMS: Knowledge of plant resource acquisition strategies is crucial for understanding the mechanisms mediating the responses of ecosystems to external nitrogen (N) input. However, few studies have considered the joint effects of above-ground (light) and below-ground (nutrient) resource acquisition strategies in regulating plant species responses to N enrichment. Here, we quantified the effects of light and non-N nutrient acquisition capacities on species relative abundance in the case of extra N input. METHODS: Based on an N-manipulation experiment in a Tibetan alpine steppe, we determined the responses of species relative abundances and light and nutrient acquisition capacities to N enrichment for two species with different resource acquisition strategies (the taller Stipa purpurea, which is colonized by arbuscular mycorrhizal fungi, and the shorter Carex stenophylloides, which has cluster roots). Structural equation models were developed to explore the relative effects of light and nutrient acquisition on species relative abundance along the N addition gradient. KEY RESULTS: We found that the relative abundance of taller S. purpurea increased with the improved light acquisition along the N addition gradient. In contrast, the shorter C. stenophylloides, with cluster roots, excelled in acquiring phosphorus (P) so as to elevate its leaf P concentration under N enrichment by producing large amounts of carboxylate exudates that mobilized moderately labile and recalcitrant soil P forms. The increased leaf P concentration of C. stenophylloides enhanced its light use efficiency and promoted its relative abundance even in the shade of taller competitors. CONCLUSIONS: Our findings highlight that the combined effects of above-ground (light) and below-ground (nutrient) resources rather than light alone (the prevailing perspective) determine the responses of grassland community structure to N enrichment.

Permafrost nitrogen status and its determinants on the Tibetan Plateau.

It had been suggested that permafrost thaw could promote frozen nitrogen (N) release and modify microbial N transformation rates, which might alter soil N availability and then regulate ecosystem functions. However, the current understanding of this issue is confined to limited observations in the Arctic permafrost region, without any systematic measurements in other permafrost regions. Based on a large-scale field investigation along a 1,000 km transect and a laboratory incubation experiment with a 15 N pool dilution approach, this study provides the comprehensive evaluation of the permafrost N status, including the available N content and related N transformation rates, across the Tibetan alpine permafrost region. In contrast to the prevailing view, our results showed that the Tibetan alpine permafrost had lower available N content and net N mineralization rate than the active layer. Moreover, the permafrost had lower gross rates of N mineralization, microbial immobilization and nitrification than the active layer. Our results also revealed that the dominant drivers of the gross N mineralization and microbial immobilization rates differed between the permafrost and the active layer, with these rates being determined by microbial properties in the permafrost while regulated by soil moisture in the active layer. In contrast, soil gross nitrification rate was consistently modulated by the soil NH 4 + content in both the permafrost and the active layer. Overall, patterns and drivers of permafrost N pools and transformation rates observed in this study offer new insights into the potential N release upon permafrost thaw and provide important clues for Earth system models to better predict permafrost biogeochemical cycles under a warming climate.

Magnitude and Drivers of Potential Methane Oxidation and Production across the Tibetan Alpine Permafrost Region.

Methane (CH4) dynamics across permafrost regions is critical in determining the magnitude and direction of permafrost carbon (C)-climate feedback. However, current studies are mainly derived from the Arctic area, with limited evidence from other permafrost regions. By combining large-scale laboratory incubation across 51 sampling sites with machine learning techniques and bootstrap analysis, here, we determined regional patterns and dominant drivers of CH4 oxidation potential in alpine steppe and meadow (CH4 sink areas) and CH4 production potential in swamp meadow (CH4 source areas) across the Tibetan alpine permafrost region. Our results showed that both CH4 oxidation potential (in alpine steppe and meadow) and CH4 production potential (in swamp meadow) exhibited large variability across various sampling sites, with the median value being 8.7, 9.6, and 11.5 ng g-1 dry soil h-1, respectively. Our results also revealed that methanotroph abundance and soil moisture were two dominant factors regulating CH4 oxidation potential, whereas CH4 production potential was mainly affected by methanogen abundance and the soil organic carbon content, with functional gene abundance acting as the best explaining variable. These results highlight the crucial role of microbes in regulating CH4 dynamics, which should be considered when predicting the permafrost C cycle under future climate scenarios.

Unimodal Response of Soil Methane Consumption to Increasing Nitrogen Additions.

Nitrogen (N) status has a great impact on methane (CH4) consumption by soils. Modeling studies predicting soil CH4 consumption assume a linear relationship between CH4 uptake and N addition rate. Here, we present evidence that a nonlinear relationship may better characterize changes in soil CH4 uptake with increasing N additions. By conducting a field experiment with eight N-input levels in a Tibetan alpine steppe, we observed a unimodal relationship; CH4 uptake increased at low to medium N levels but declined at high N levels. Environmental and microbial properties jointly determined this response pattern. The generality of the unimodal trend was further validated by two independent analyses: (i) we examined soil CH4 uptake across at least five N-input levels in upland ecosystems across China. A unimodal CH4 uptake-N addition rate relationship was observed in 3 out of 4 cases; and (ii) we performed a meta-analysis to explore the N-induced changes in soil CH4 uptake with increasing N additions across global upland ecosystems. Results showed that the changes in CH4 uptake exhibited a quadratic correlation with N addition rate. Overall, we suggest that the unimodal relationship should be considered in biogeochemistry models for accurately predicting soil CH4 consumption under global N enrichment.

Changes in Methane Flux along a Permafrost Thaw Sequence on the Tibetan Plateau.

Permafrost thaw alters the physical and environmental conditions of soil and may thus cause a positive feedback to climate warming through increased methane emissions. However, the current knowledge of methane emissions following thermokarst development is primarily based on expanding lakes and wetlands, with upland thermokarst being studied less often. In this study, we monitored the methane emissions during the peak growing seasons of two consecutive years along a thaw sequence within a thermo-erosion gully in a Tibetan swamp meadow. Both years had consistent results, with the early and midthaw stages (3 to 12 years since thaw) exhibiting low methane emissions that were similar to those in the undisturbed meadow, while the emissions from the late thaw stage (20 years since thaw) were 3.5 times higher. Our results also showed that the soil water-filled pore space, rather than the soil moisture per se, in combination with the sand content, were the main factors that caused increased methane emissions. These findings differ from the traditional view that upland thermokarst could reduce methane emissions owing to the improvement of drainage conditions, suggesting that upland thermokarst development does not always result in a decrease in methane emissions.

Magnitude and Pathways of Increased Nitrous Oxide Emissions from Uplands Following Permafrost Thaw.

Permafrost thawing may release nitrous oxide (N2O) due to large N storage in cold environments. However, N2O emissions from permafrost regions have received little attention to date, particularly with respect to the underlying microbial mechanisms. We examined the magnitude of N2O fluxes following upland thermokarst formation along a 20-year thaw sequence within a thermo-erosion gully in a Tibetan swamp meadow. We also determined the importance of environmental factors and the related microbial functional gene abundance. Our results showed that permafrost thawing led to a mass release of N2O in recently collapsed sites (3 years ago), particularly in exposed soil patches, which presented post-thaw emission rates equivalent to those from agricultural and tropical soils. In addition to abiotic factors, soil microorganisms exerted significant effects on the variability in the N2O emissions along the thaw sequence and between vegetated and exposed patches. Overall, our results demonstrate that upland thermokarst formation can lead to enhanced N2O emissions, and that the global warming potential (GWP) of N2O at the thermokarst sites can reach 60% of the GWP of CH4 (vs ∼6% in control sites), highlighting the potentially strong noncarbon (C) feedback to climate warming in permafrost regions.

Warming effects on permafrost ecosystem carbon fluxes associated with plant nutrients.

Large uncertainties exist in carbon (C)-climate feedback in permafrost regions, partly due to an insufficient understanding of warming effects on nutrient availabilities and their subsequent impacts on vegetation C sequestration. Although a warming climate may promote a substantial release of soil C to the atmosphere, a warming-induced increase in soil nutrient availability may enhance plant productivity, thus offsetting C loss from microbial respiration. Here, we present evidence that the positive temperature effect on carbon dioxide (CO2 ) fluxes may be weakened by reduced plant nitrogen (N) and phosphorous (P) concentrations in a Tibetan permafrost ecosystem. Although experimental warming initially enhanced ecosystem CO2 uptake, the increased rate disappeared after the period of peak plant growth during the early growing season, even though soil moisture was not a limiting factor in this swamp meadow ecosystem. We observed that warming did not significantly affect soil extractable N or P during the period of peak growth, but decreased both N and P concentrations in the leaves of dominant plant species, likely caused by accelerated plant senescence in the warmed plots. The attenuated warming effect on CO2 assimilation during the late growing season was associated with lowered leaf N and P concentrations. These findings suggest that warming-mediated nutrient changes may not always benefit ecosystem C uptake in permafrost regions, making our ability to predict the C balance in these warming-sensitive ecosystems more challenging than previously thought.

Linkages of plant stoichiometry to ecosystem production and carbon fluxes with increasing nitrogen inputs in an alpine steppe.

Unprecedented levels of nitrogen (N) have entered terrestrial ecosystems over the past century, which substantially influences the carbon (C) exchange between the atmosphere and biosphere. Temperature and moisture are generally regarded as the major controllers over the N effects on ecosystem C uptake and release. N-phosphorous (P) stoichiometry regulates the growth and metabolisms of plants and soil organisms, thereby affecting many ecosystem C processes. However, it remains unclear how the N-induced shift in the plant N:P ratio affects ecosystem production and C fluxes and its relative importance. We conducted a field manipulative experiment with eight N addition levels in a Tibetan alpine steppe and assessed the influences of N on aboveground net primary production (ANPP), gross ecosystem productivity (GEP), ecosystem respiration (ER), and net ecosystem exchange (NEE); we used linear mixed-effects models to further determine the relative contributions of various factors to the N-induced changes in these parameters. Our results showed that the ANPP, GEP, ER, and NEE all exhibited nonlinear responses to increasing N additions. Further analysis demonstrated that the plant N:P ratio played a dominate role in shaping these C exchange processes. There was a positive relationship between the N-induced changes in ANPP (ΔANPP) and the plant N:P ratio (ΔN:P), whereas the ΔGEP, ΔER, and ΔNEE exhibited quadratic correlations with the ΔN:P. In contrast, soil temperature and moisture were only secondary predictors for the changes in ecosystem production and C fluxes along the N addition gradient. These findings highlight the importance of plant N:P ratio in regulating ecosystem C exchange, which is crucial for improving our understanding of C cycles under the scenarios of global N enrichment.

Dual roles of microbes in mediating soil carbon dynamics in response to warming.

Understanding the alterations in soil microbial communities in response to climate warming and their controls over soil carbon (C) processes is crucial for projecting permafrost C-climate feedback. However, previous studies have mainly focused on microorganism-mediated soil C release, and little is known about whether and how climate warming affects microbial anabolism and the subsequent C input in permafrost regions. Here, based on a more than half-decade of in situ warming experiment, we show that compared with ambient control, warming significantly reduces microbial C use efficiency and enhances microbial network complexity, which promotes soil heterotrophic respiration. Meanwhile, microbial necromass markedly accumulates under warming likely due to preferential microbial decomposition of plant-derived C, further leading to the increase in mineral-associated organic C. Altogether, these results demonstrate dual roles of microbes in affecting soil C release and stabilization, implying that permafrost C-climate feedback would weaken over time with dampened response of microbial respiration and increased proportion of stable C pool.

Metagenomic insights into microbial community structure and metabolism in alpine permafrost on the Tibetan Plateau.

Permafrost, characterized by its frozen soil, serves as a unique habitat for diverse microorganisms. Understanding these microbial communities is crucial for predicting the response of permafrost ecosystems to climate change. However, large-scale evidence regarding stratigraphic variations in microbial profiles remains limited. Here, we analyze microbial community structure and functional potential based on 16S rRNA gene amplicon sequencing and metagenomic data obtained from an ∼1000 km permafrost transect on the Tibetan Plateau. We find that microbial alpha diversity declines but beta diversity increases down the soil profile. Microbial assemblages are primarily governed by dispersal limitation and drift, with the importance of drift decreasing but that of dispersal limitation increasing with soil depth. Moreover, genes related to reduction reactions (e.g., ferric iron reduction, dissimilatory nitrate reduction, and denitrification) are enriched in the subsurface and permafrost layers. In addition, microbial groups involved in alternative electron accepting processes are more diverse and contribute highly to community-level metabolic profiles in the subsurface and permafrost layers, likely reflecting the lower redox potential and more complicated trophic strategies for microorganisms in deeper soils. Overall, these findings provide comprehensive insights into large-scale stratigraphic profiles of microbial community structure and functional potentials in permafrost regions.

Progressive nitrogen limitation across the Tibetan alpine permafrost region.

The ecosystem carbon (C) balance in permafrost regions, which has a global significance in understanding the terrestrial C-climate feedback, is significantly regulated by nitrogen (N) dynamics. However, our knowledge on temporal changes in vegetation N limitation (i.e., the supply of N relative to plant N demand) in permafrost ecosystems is still limited. Based on the combination of isotopic observations derived from a re-sampling campaign along a ~3000 km transect and simulations obtained from a process-based biogeochemical model, here we detect changes in ecosystem N cycle across the Tibetan alpine permafrost region over the past decade. We find that vegetation N limitation becomes stronger despite the increased available N production. The enhanced N limitation on vegetation growth is driven by the joint effects of elevated plant N demand and gaseous N loss. These findings suggest that N would constrain the future trajectory of ecosystem C cycle in this alpine permafrost region.