Gasdermin D (GSDMD) executes the cell death program of pyroptosis by assembling into oligomers that permeabilize the plasma membrane. Here, by single-molecule imaging, we elucidate the yet unclear mechanism of Gasdermin D pore assembly and the role of cysteine residues in GSDMD oligomerization. We show that GSDMD preassembles at the membrane into dimeric and trimeric building blocks that can either be inserted into the membrane, or further assemble into higher-order oligomers prior to insertion into the membrane. The GSDMD residues Cys39, Cys57, and Cys192 are the only relevant cysteines involved in GSDMD oligomerization. S-palmitoylation of Cys192, combined with the presence of negatively-charged lipids, controls GSDMD membrane targeting. Simultaneous Cys39/57/192-to-alanine (Ala) mutations, but not Ala mutations of Cys192 or the Cys39/57 pair individually, completely abolish GSDMD insertion into artificial membranes as well as into the plasma membrane. Finally, either Cys192 or the Cys39/Cys57 pair are sufficient to enable formation of GSDMD dimers/trimers, but they are all required for functional higher-order oligomer formation. Overall, our study unveils a cooperative role of Cys192 palmitoylation-mediated membrane binding and Cys39/57/192-mediated oligomerization in GSDMD pore assembly. This study supports a model in which Gasdermin D oligomerization relies on a two-step mechanism mediated by specific cysteine residues.

In severely phosphorus (P)-impoverished environments, plants have evolved to use P very efficiently. Yet, it is unclear how P allocation in leaves contributes to their photosynthetic P-use efficiency (PPUE) and position along the leaf economics spectrum (LES). We address this question in 10 species of Banksia and Hakea, two highly P-efficient Proteaceae genera. We characterised traits in leaves of Banksia and Hakea associated with the LES: leaf mass per area, light-saturated photosynthetic rates, P and nitrogen concentrations, and PPUE. We also determined leaf P partitioning to five biochemical fractions (lipid, nucleic acid, metabolite, inorganic and residual P) and their possible association with the LES. For both genera, PPUE was negatively correlated with fractional allocation of P to lipids, but positively correlated with that to metabolites. For Banksia only, PPUE was negatively correlated with residual P, highlighting a strategy contrasting to that of Hakea. Phosphorus-allocation patterns significantly explained PPUE but were not linked to the resource acquisition vs resource conservation gradient defined by the LES. We conclude that distinct P-allocation patterns enable species from different genera to achieve high PPUE and discuss the implications of different P investments. We surmise that different LES axes representing different ecological strategies coexist in extremely P-impoverished environments.

Traditionally viewed as a fixed and homogeneous machinery for protein synthesis, the ribosome is increasingly recognized for its heterogeneity, as indicated by emerging studies highlighting the functional relevance of specialized ribosomes. However, whether ribosome heterogeneity is merely an outcome limited to specific conditions or a pervasive cellular phenomenon remains unclear, and existing evidence on the extensive existence of ribosome heterogeneity is scant. Here, we leveraged existing proteomic data and employed ribosome ratio-omics (RibosomeR), which comprehensively analyzes ribosome protein stoichiometry across various biological samples exhibiting distinct functions, developmental stages, and pathological states. Using the 80S monosome proteomic data, RibosomeR analysis unveils significant ribosome heterogeneity across different tissues, including fat, spleen, liver, kidney, heart, and skeletal muscles. Furthermore, examination of testes at various stages of spermatogenesis reveals distinct RibosomeR signatures during tissue development. Analysis of the whole cell proteomic data finds that RibosomeR undergoes dynamic changes during in vitro neuronal maturation, indicating functional associations with specific molecular aspects of neurodevelopment. In pathological contexts, RibosomeR signatures in gastric tumors demonstrate functional links to pathways associated with tumorigenesis. Additionally, dynamic alterations in RibosomeR are observed in macrophages following immune challenges. Collectively, our investigation across a diverse array of biological samples underscores the presence of ribosome heterogeneity, while previous studies observed functional aspects of ribosome specialization, in cellular function, development, and disease. The RibosomeR barcode serves as a valuable tool for elucidating these complexities.

Soil enzyme activities are pivotal for diverse biochemical processes and are sensitive to land use changes. They can indicate soil microbial nutrient limitations. Nonetheless, the mechanism governing the response of soil microbial nutrient limitation to land use alterations in coastal regions remains elusive. We assessed soil nutrients, microbial biomass, and extracellular enzyme activities across various land use types-natural (wasteland and woodland) and agricultural (farmland and orchard)-in the Hangzhou Bay area, China. All four land use types experience co-limitation by carbon (C) and phosphorus (P). However, the extent of microbial resource limitations varies among them. Long-term agricultural practices diminish microbial C and P limitations in farmland and orchard soils compared to natural soils, as evidenced by lower ecoenzymatic C:N ratios and vector lengths, alongside higher microbial carbon use efficiency (CUE). Soil nutrient stoichiometric ratios and CUE are primary factors influencing microbial C and P limitations. Thus, fostering appropriate land use and management practices proves imperative to regulate soil nutrient cycles and foster the sustainable management of coastal areas.

CX-5461, also known as pidnarulex, is a strong G4 stabilizer and has received FDA fast-track designation for BRCA1- and BRCA2- mutated cancers. However, quantitative measurements of the unfolding rates of CX-5461-G4 complexes which are important for the regulation function of G4s, remain lacking. Here, we employ single-molecule magnetic tweezers to measure the unfolding force distributions of c-MYC G4s in the presence of different concentrations of CX-5461. The unfolding force distributions exhibit three discrete levels of unfolding force peaks, corresponding to three binding modes. In combination with a fluorescent quenching assay and molecular docking to previously reported ligand-c-MYC G4 structure, we assigned the ~69 pN peak corresponding to the 1:1 (ligand:G4) complex where CX-5461 binds at the G4\'s 5\'-end. The ~84 pN peak is attributed to the 2:1 complex where CX-5461 occupies both the 5\' and 3\'. Furthermore, using the Bell-Arrhenius model to fit the unfolding force distributions, we determined the zero-force unfolding rates of 1:1, and 2:1 complexes to be (2.4 ± 0.9) × 10-8 s-1 and (1.4 ± 1.0) × 10-9 s-1 respectively. These findings provide valuable insights for the development of G4-targeted ligands to combat c-MYC-driven cancers.

Microorganisms grow despite imbalances in the availability of nutrients and energy. The biochemical and elemental adjustments that bacteria employ to sustain growth when these resources are suboptimal are not well understood. We assessed how Pseudomonas putida KT2440 adjusts its physiology at differing dilution rates (to approximate growth rates) in response to carbon (C), nitrogen (N), and phosphorus (P) stress using chemostats. Cellular elemental and biomolecular pools were variable in response to different limiting resources at a slow dilution rate of 0.12 h-1, but these pools were more similar across treatments at a faster rate of 0.48 h-1. At slow dilution rates, limitation by P and C appeared to alter cell growth efficiencies as reflected by changes in cellular C quotas and rates of oxygen consumption, both of which were highest under P- and lowest under C- stress. Underlying these phenotypic changes was differential gene expression of terminal oxidases used for ATP generation that allows for increased energy generation efficiency. In all treatments under fast dilution rates, KT2440 formed aggregates and biofilms, a physiological response that hindered an accurate assessment of growth rate, but which could serve as a mechanism that allows cells to remain in conditions where growth is favorable. Our findings highlight the ways that microorganisms dynamically adjust their physiology under different resource supply conditions, with distinct mechanisms depending on the limiting resource at slow growth and convergence toward an aggregative phenotype with similar compositions under conditions that attempt to force fast growth.
OBJECTIVE: All organisms experience suboptimal growth conditions due to low nutrient and energy availability. Their ability to survive and reproduce under such conditions determines their evolutionary fitness. By imposing suboptimal resource ratios under different dilution rates on the model organism Pseudomonas putida KT2440, we show that this bacterium dynamically adjusts its elemental composition, morphology, pools of biomolecules, and levels of gene expression. By examining the ability of bacteria to respond to C:N:P imbalance, we can begin to understand how stoichiometric flexibility manifests at the cellular level and impacts the flow of energy and elements through ecosystems.

Chilled and cut chicken is preferred by consumers for its safeness and readiness to cook. To evaluate the quality characteristics of various chilled chicken products, differences in volatile organic components (VOCs) of six different cut parts (breast, back, leg, heart, liver, and gizzard) of Lueyang black chicken were characterized through gas chromatography-ion mobility spectroscopy (GC-IMS) combined with stoichiometry. A total of 54 peaks in the signal of VOCs were detected by GC-IMS, and 43 VOCs were identified by qualitative analysis. There were 22 aldehydes (20.66-54.07%), 8 ketones (25.74-62.87%), 9 alcohols (4.17-14.69%), 1 ether (0.18-2.22%), 2 esters (0.43-1.54%), and 1 furan (0.13-0.52%), in which aldehydes, ketones, and alcohols were the main categories. Among the six cut parts, the relative content of aldehydes (54.07%) was the highest in the gizzard, and the relative content of ketones (62.87%) was the highest in the heart. Meanwhile, the relative content of alcohols (14.69%) was the highest in the liver. Based on a stable and reliable predictive model established by orthogonal partial least squares-discriminant analysis (OPLS-DA), 3-hydroxy-2-butanone (monomer and dimer), acetone, 2-butanone monomer, hexanal (monomer and dimer), isopentyl alcohol monomer, and n-hexanol monomer were picked out as characteristic VOCs based on variable importance in projection (VIP value > 1.0, p < 0.05). Principal component analysis (PCA) and the clustering heatmap indicated that the characteristic VOCs could effectively distinguish the six cut parts of Lueyang black chicken. The specific VOCs responsible for flavor differences among six different cut parts of Lueyang black chicken were hexanal (monomer and dimer) for the gizzard, 2-butanone monomer and hexanal dimer for the breast, hexanal monomer for the back, 3-hydroxy-2-butanone monomer for the leg, 3-hydroxy-2-butanone (monomer and dimer) for the heart, and acetone and isopentyl alcohol monomer for the liver. These findings could reveal references for quality assessment and development of chilled products related to different cut parts of Lueyang black chicken in the future.

Human activities have increased nitrogen (N) and phosphorus (P) inputs to the Yellow River Delta and the supply level of N and P affects plant growth as well as ecosystem structure and function directly. However, the root growth, stoichiometry, and antioxidant system of plants in response to N and P additions, especially for herbaceous halophyte in the Yellow River Delta (YRD), remain unknown. A field experiment with N addition (0, 5, 15, and 45 g N m-2 yr-1, respectively) as the main plot, and P addition (0 and 1 g N m-2 yr-1, respectively) as the subplot, was carried out with a split-plot design to investigate the effects on the root morphology, stoichiometry, and antioxidant system of Suaeda salsa. The results showed that N addition significantly increased the above-ground and root biomass as well as shoot-root ratio of S. salsa, which had a significant interaction with P addition. The highest biomass was found in the treatment with 45 g N m-2 yr-1 combined with P addition. N addition significantly increased TN content and decreased C:N ratio of root, while P addition significantly increased TP content and decreased C:P ratio. The main root length (MRL), total root length (TRL), specific root length (SRL), and root tissue density (RTD) of S. salsa root were significantly affected by N addition and P addition, as well as their interaction. The treatments with or without P addition at the 45 g N m-2 yr-1 of N addition significantly increased the superoxide dismutase (SOD), peroxidase (POD), catalase (CAT) activities and soluble protein content of roots, decreased malondialdehyde (MDA) content. And there was a significant interaction between the N and P addition on SOD activity. Therefore, N and P additions could improve the growth of S. salsa by altering the root morphology, increasing the root nutrient content, and stimulating antioxidant system.

Phytoplankton Chl:C:N:P ratios are important from both an ecological and a biogeochemical perspective. We show that these elemental ratios can be represented by a phytoplankton physiological model of low complexity that includes major cellular macromolecular pools. In particular, our model resolves time-dependent intracellular pools of chlorophyll, proteins, nucleic acids, carbohydrates/lipids, and N and P storage. Batch culture data for two diatom and two prasinophyte species are used to constrain parameters that represent specific allocation traits and strategies. A key novelty is the simultaneous estimation of physiological parameters for two phytoplankton groups of such different sizes. The number of free parameters is reduced by assuming (i) allometric scaling for maximum uptake rates, (ii) shared half-saturation constants for synthesis of functional macromolecules, (iii) shared exudation rates of functional macromolecules across the species. The rationale behind this assumption is that across the different species, the same or similar processes, enzymes, and metabolites play a role in key physiological processes. For the turnover numbers of macromolecular synthesis and storage exudation rates, differences between diatoms and prasinophytes need to be taken into account to obtain a good fit. Our model fits suggest that the parameters related to storage dynamics dominate the differences in the C:N:P ratios between the different phytoplankton groups. Since descriptions of storage dynamics are still incomplete and imprecise, predictions of C:N:P ratios by phytoplankton models likely have a large uncertainty.

The long-term stability of perovskite solar cells (PSCs) remains a bottleneck for commercialization. While studies on the stoichiometry and morphology of PSCs with regard to performance are prevalent, understanding the influence of these factors on their long-term stability is lacking. In this work, we evaluate the impact of stoichiometry and morphology on the long-term stability of cesium formamidinium-based PSCs. We demonstrate that the lead iodide (PbI2) to formamidinium iodide (FAI) ratio influences stability under various stress factors (elevated temperature and light). A high molar ratio (PbI2/FAI > 1.1) in the perovskite precursor displays drastic degradation under ISOS-L1 (100 mW/cm2, 25 °C, maximum power point tracking) conditions. However, postdegradation analysis contradicts these results. Devices with PbI2/FAI ≤ 1.1 are stable under light, but intermittent current density-voltage characterizations indicate that device performance decreases during storage in the dark. Migration of iodide (I-) ions to the electron-transport layer (ETL) and iodine vacancies (VI-+) to the hole-transport layer (HTL) forms localized shunts in the absorber layer. Pinhole formation, surrounded by FA+-rich regions, explains the extent of damage in comparably aged films. In summary, this work emphasizes the importance of reporting stability under different stress conditions, coupled with postdegradation and dark recovery analyses of PSCs to better understand the complexities of perovskite instability under real-life conditions such as expected during outdoor operation.