Integration of Hybridization Chain Reaction and Protein-Binding Amplification for Long-Term Imaging of Intracellular mRNA: Avoiding Signal Fluctuation.

Amplified nanoprobes based on hybridization chain reaction (HCR) have been widely developed for the detection of intracellular low abundance mRNA. However, the formed chain-like assembly decorated with fluorophore would be degraded rapidly by endogenous enzyme, resulting in failure of the long-term fluorescence imaging. To address this issue, herein, a composite signal-amplifying strategy that integrates HCR into protein-binding signal amplification (HPSA) was communicated for the in situ imaging of mRNA by avoiding signal fluctuation. Different from conventional HCR-based nanoprobes (HCR-nanoprobe), the HCR was used as the signal-triggered mode and the amplifying signal generated from in situ fluorophore-protein binding in cells, which can maintain high stability of the signal for a long time. As a proof-of-principle, a nanobeacon based on HPSA (HPSA-nanobeacon) was constructed to detect TK1 mRNA. Taking advantage of the double signal-amplifying mode, the endogenous TK1 mRNA was sensitively detected and the fluorescence signal was maintained for more than 8 h in HepG2 cells. The attempt in this work provides a new option to the current signal-amplifying strategy for sensing nucleic acid targets with high stability, significantly enhancing the acquisition of intracellular molecular information.

Seeds exhibit the most stable elemental composition with nitrogen addition in an Inner Mongolian grassland.

Variation in biomass elemental composition of grassland plants may have important implications for ecosystem functioning in response to global change. However, relevant studies have mostly focused on variation of nitrogen (N) and phosphorus (P) concentrations in plant leaves, while few studies have evaluated other elements and plant organs of grassland species. Here, we examined the effects of N addition on multi-element concentrations, and analyzed their patterns across different organs (leaf, stem, root and seed) of five plant species in a steppe community of the Inner Mongolian grassland. Our results showed that seeds exhibited the most stable elemental composition with N addition, and that manganese (Mn) and iron (Fe) concentrations were substantially more variable than macro-elements in response to N addition. In particular, we identified a set of significant negative relationships between elemental concentrations and their corresponding CVs (coefficients of variation) for all plant organs as a whole and for each individual organ. We further found that changes in soil pH and the availability of soil nutrients contributed mostly to variation in the biomass elemental composition of major plants in this community. These findings are important for accurately assessing the effects of N deposition on the biochemical cycling of nutrient elements in grassland ecosystems, and provide critical clues for developing effective approaches to adaptively managing grassland resources as well as mitigating the impact of global change on the dryland ecosystems in the Mongolia Plateau.

Fibroblast activation protein-sensitive polymeric nanobeacon for early diagnosis of renal fibrosis.

Early diagnosis and treatment of renal fibrosis (RF) significantly affect the clinical outcomes of chronic kidney diseases (CKDs). As the typical fibrotic ailment, RF is characterized by remodeling of the extracellular matrix, and the activation of fibroblast activation protein (FAP) plays a crucial role in the mediation of extracellular matrix protein degradation. Therefore, FAP can serve as a biomarker for RF. However, up to now, no effective tools have been reported to diagnose early-stage RF via detecting FAP. In this work, a polymeric nanobeacon integrating an FAP-sensitive amphiphilic polymer and fluorophores was proposed, which was used to diagnose early RF by sensing FAP. The FAP can be detected in the range of 0 to 200 ng/mL with a detection limit of 0.132 ng/mL. Furthermore, the fluorescence imaging results demonstrate that the polymeric nanobeacon can sensitively image fibrotic kidneys in mice with unilateral ureteral occlusion (UUO), suggesting its potential for early RF diagnosis and guidance of FAP-targeted treatments. Importantly, when employed alongside with non-invasive diagnostic techniques like magnetic resonance imaging (MRI) and serological tests, this nanobeacon exhibits excellent biocompatibility, low biological toxicity, and sustained imaging capabilities, making it a suitable fluorescent tool for diagnosing various FAP-related fibrotic conditions. To our knowledge, this study represents the first attempt to image RF in early stage by detecting FAP, offering a promising fluorescent molecular tool for diagnosing various FAP-associated diseases in the future.

Soil Acidification Under Long-Term N Addition Decreases the Diversity of Soil Bacteria and Fungi and Changes Their Community Composition in a Semiarid Grassland.

Soil microorganisms play key roles in terrestrial biogeochemical cycles and ecosystem functions. However, few studies address how long-term nitrogen (N) addition gradients impact soil bacterial and fungal diversity and community composition simultaneously. Here, we investigated soil bacterial and fungal diversity and community composition based on a long-term (17 years) N addition gradient experiment (six levels: 0, 2, 4, 8, 16, 32 gN m-2 year-1) in temperate grassland, using the high-throughput Illumina MiSeq sequencing. Results showed that both soil bacterial and fungal alpha diversity responded nonlinearly to the N input gradient and reduced drastically when the N addition rate reached 32 g N m-2 year-1. The relative abundance of soil bacterial phyla Proteobacteria increased and Acidobacteria decreased significantly with increasing N level. In addition, the relative abundance of bacterial functional groups associated with aerobic ammonia oxidation, aerobic nitrite oxidation, nitrification, respiration of sulfate and sulfur compounds, and chitinolysis significantly decreased under the highest N addition treatment. For soil fungi, the relative abundance of Ascomycota increased linearly along the N enrichment gradient. These results suggest that changes in soil microbial community composition under elevated N do not always support the copiotrophic-oligotrophic hypothesis, and some certain functional bacteria would not simply be controlled by soil nutrients. Further analysis illustrated that reduced soil pH under N addition was the main factor driving variations in soil microbial diversity and community structure in this grassland. Our findings highlight the consistently nonlinear responses of soil bacterial and fungal diversity to increasing N input and the significant effects of soil acidification on soil microbial communities, which can be helpful for the prediction of underground ecosystem processes in light of future rising N deposition.

Spatiotemporal patterns and drivers of methane uptake across a climate transect in Inner Mongolia Steppe.

Steppe soils are important biological sinks for atmospheric methane (CH4), but the strength of CH4 uptake remains uncertain due to large spatiotemporal variation and the lack of in situ measurements at regional scale. Here, we report the seasonal and spatial patterns of CH4 uptake across a 1200 km transect in arid and semi-arid steppe ecosystems in Inner Mongolia, ranging from meadow steppe in the east plain to typical and desert steppes on the west plateau. In general, seasonal patterns of CH4 uptake were site specific, with unimodal seasonal curves in meadow and typical steppes and a decreasing seasonal trend in desert steppe. Soil moisture was the dominant factor explaining the seasonal patterns of CH4 uptake, and CH4 uptake rate decreased with an increase in soil moisture. Across the transect, CH4 uptake showed a skewed unimodal spatial pattern, with the peak rate observed in the typical steppe sites and with generally higher uptake rates in the west plateau than in the east plain. Soil moisture, together with soil temperature, soil total carbon, and aboveground plant biomass, were the main drivers of the regional patterns of CH4 uptake rate. These findings are important for model development to more precisely estimate the soil CH4 sink capacity in arid and semi-arid regions.

Response of soil methane uptake to simulated nitrogen deposition and grazing management across three types of steppe in Inner Mongolia, China.

The response of soil methane (CH4) uptake to increased nitrogen (N) deposition and grazing management was studied in three types of steppe (i.e., meadow steppe, typical steppe, and desert steppe) in Inner Mongolia, China. The experiment was designed with four simulated N deposition rates such as 0, 50, 100, and 200kgNha-1, respectively, under grazed and fenced management treatments. Results showed that the investigated steppes were significant sinks for CH4, with an uptake flux of 1.12-3.36kgha-1 over the grass growing season and that the magnitude of CH4 uptake significantly (P<0.05) decreased with increasing N deposition rates. The soil CH4 uptake rates were highest in the desert steppe, moderate in the typical steppe, and lowest in the meadow steppe. Compared with grazed plots, fencing increased the CH4 uptake by 4.7-40.2% with a mean value of 20.2% across the three different steppe types. The responses of soil CH4 uptake to N deposition in the continental steppe varied depending on the N deposition rate, steppe type, and grazing management. A significantly positive correlation between CH4 uptake and soil temperature was found in this study, whereas no significant relationship between soil moisture and CH4 uptake occurred. Our results may contribute to the improvement of model parameterization for simulating biosphere-atmosphere CH4 exchange processes and for evaluating the climate change feedback on CH4 soil uptake.

Effects of water and nitrogen addition on ecosystem respiration across three types of steppe: The role of plant and microbial biomass.

Evaluating the regional variation of ecosystem respiration (Reco) in its response to the changes of soil water and nitrogen (N) availability is crucial for fully understanding ecosystem carbon (C) exchange and its feedbacks to global changes. Here, we examined the responses of Reco, plant community aboveground biomass (AB), microbial biomass carbon (MBC) and soil moisture (SM) to water and N addition, using intact soil monoliths from three different temperate steppes along a precipitation gradient, including meadow steppe, typical steppe, and desert steppe in northern China. We found that the meadow steppe held the highest value of Reco. Water addition significantly enhanced Reco while N addition had no effect on Reco in all three ecosystems. The response of Reco in the typical steppe was more sensitive than the other two ecosystems. The changes of plant community AB exhibited a much stronger explanatory power than that of MBC for Reco in the typical steppe. In contrast, MBC was the dominant factor explaining the variation of Reco in the desert steppe and the meadow steppe. These findings contribute to our understanding of regional patterns of ecosystem C exchange under scenarios of global changes and highlight the importance of water availability in regulating ecosystem processes in temperate steppe grasslands.

Seasonal and Spatial Variations of Bulk Nitrogen Deposition and the Impacts on the Carbon Cycle in the Arid/Semiarid Grassland of Inner Mongolia, China.

Atmospheric nitrogen (N) deposition is an important component that affects the structure and function of different terrestrial ecosystem worldwide. However, much uncertainty still remains concerning the magnitude of N deposition on grassland ecosystem in China. To study the spatial and temporal patterns of bulk N deposition, the levels of N (NH4+-N and NO3--N) concentration in rainfall were measured at 12 sites across a 1200 km grassland transect in Inner Mongolia, China, and the respective N deposition rates were estimated. The inorganic N deposition rates ranged from 4.53 kg N ha-1 to 12.21 kg N ha-1 with a mean value of 8.07 kg N ha-1 during the entire growing season, decreasing steadily from the eastern to the western regions. Inorganic N deposition occurred mainly in July and August across meadow steppe, typical steppe, and desert steppe, which corresponded to the seasonal distribution of mean annual precipitation. A positive relationship was found between inorganic N deposition and mean annual precipitation (R2 = 0.54 ~ 0.72, P < 0.0001) across the grassland transect. Annual estimation of inorganic N deposition was 0.67 Pg yr-1 in Inner Mongolia, China based on the correlation between N deposition rates and precipitation. N deposition was an important factor controlling aboveground biomass and ecosystem respiration, but has no effect on root biomass and soil respiration. We must clarify that we used the bulk deposition samplers during the entire sampling process and estimated the dissolved NH4+-N and NO3--N deposition rates during the entire growing season. Long-term N deposition monitoring networks should be constructed to study the patterns of N deposition and its potential effect on grassland ecosystem, considering various N species, i.e., gaseous N, particle N, and wet N deposition.

Effects of elevated CO2 and N addition on growth and N2 fixation of a legume subshrub (Caragana microphylla Lam.) in temperate grassland in China.

It is well demonstrated that the responses of plants to elevated atmospheric CO(2) concentration are species-specific and dependent on environmental conditions. We investigated the responses of a subshrub legume species, Caragana microphylla Lam., to elevated CO(2) and nitrogen (N) addition using open-top chambers in a semiarid temperate grassland in northern China for three years. Measured variables include leaf photosynthetic rate, shoot biomass, root biomass, symbiotic nitrogenase activity, and leaf N content. Symbiotic nitrogenase activity was determined by the C(2)H(2) reduction method. Elevated CO(2) enhanced photosynthesis and shoot biomass by 83% and 25%, respectively, and the enhancement of shoot biomass was significant only at a high N concentration. In addition, the photosynthetic capacity of C. microphylla did not show down-regulation under elevated CO(2). Elevated CO(2) had no significant effect on root biomass, symbiotic nitrogenase activity and leaf N content. Under elevated CO(2), N addition stimulated photosynthesis and shoot biomass. By contrast, N addition strongly inhibited symbiotic nitrogenase activity and slightly increased leaf N content of C. microphylla under both CO(2) levels, and had no significant effect on root biomass. The effect of elevated CO(2) and N addition on C. microphylla did not show interannual variation, except for the effect of N addition on leaf N content. These results indicate that shoot growth of C. microphylla is more sensitive to elevated CO(2) than is root growth. The stimulation of shoot growth of C. microphylla under elevated CO(2) or N addition is not associated with changes in N(2)-fixation. Additionally, elevated CO(2) and N addition interacted to affect shoot growth of C. microphylla with a stimulatory effect occurring only under combination of these two factors.