High-performance peptide biosensor based on unified structure of lotus silk.

The sensitive materials of current gas sensors are fabricated on planar substrates, significantly limiting the quantity of sensitive material available on the sensor and the complete exposure of the sensitive material to the target gas. In this work, we harnessed the finest, resilient, naturally degradable, and low-cost lotus silk derived from plant fibers, to fabricate a high-performance bio-sensor for toxic and harmful gas detection, employing peptides with full surface connectivity. The proposed approach to fabricate gas sensors eliminated the need for substrates and electrodes. To ascertain the effectiveness and versatility of the sensors created via this method, sensors for three distinct representative gases (isoamyl alcohol, 4-vinylanisole, and benzene) were prepared and characterized. These sensors surpassed reported detection limits by at least one order of magnitude. The inherent pliancy of lotus silk imparts adaptability to the sensor architecture, facilitating the realization of 1D, 2D, or 3D configurations, all while upholding consistent performance characteristics. This innovative sensor paradigm, grounded in lotus silk, represents great potential toward the advancement of highly proficient bio gas sensors and associated applications.

De novo biosynthesis of the hops bioactive flavonoid xanthohumol in yeast.

The flavonoid xanthohumol is an important flavor substance in the brewing industry that has a wide variety of bioactivities. However, its unstable structure results in its low content in beer. Microbial biosynthesis is considered a sustainable and economically viable alternative. Here, we harness the yeast Saccharomyces cerevisiae for the de novo biosynthesis of xanthohumol from glucose by balancing the three parallel biosynthetic pathways, prenyltransferase engineering, enhancing precursor supply, constructing enzyme fusion, and peroxisomal engineering. These strategies improve the production of the key xanthohumol precursor demethylxanthohumol (DMX) by 83-fold and achieve the de novo biosynthesis of xanthohumol in yeast. We also reveal that prenylation is the key limiting step in DMX biosynthesis and develop tailored metabolic regulation strategies to enhance the DMAPP availability and prenylation efficiency. Our work provides feasible approaches for systematically engineering yeast cell factories for the de novo biosynthesis of complex natural products.

The ERF transcription factor LTF1 activates DIR1 to control stereoselective synthesis of antiviral lignans and stress defense in Isatis indigotica roots.

Lignans are a powerful weapon for plants to resist stresses and have diverse bioactive functions to protect human health. Elucidating the mechanisms of stereoselective biosynthesis and response to stresses of lignans is important for the guidance of plant improvement. Here, we identified the complete pathway to stereoselectively synthesize antiviral (-)-lariciresinol glucosides in Isatis indigotica roots, which consists of three-step sequential stereoselective enzymes DIR1/2, PLR, and UGT71B2. DIR1 was further identified as the key gene in respoJanuary 2024nse to stresses and was able to trigger stress defenses by mediating the elevation in lignan content. Mechanistically, the phytohormone-responsive ERF transcription factor LTF1 colocalized with DIR1 in the cell periphery of the vascular regions in mature roots and helped resist biotic and abiotic stresses by directly regulating the expression of DIR1. These systematic results suggest that DIR1 as the first common step of the lignan pathway cooperates with PLR and UGT71B2 to stereoselectively synthesize (-)-lariciresinol derived antiviral lignans in I. indigotica roots and is also a part of the LTF1-mediated regulatory network to resist stresses. In conclusion, the LTF1-DIR1 module is an ideal engineering target to improve plant Defenses while increasing the content of valuable lignans in plants.

p32 regulates glycometabolism and TCA cycle to inhibit ccRCC progression via copper-induced DLAT lipoylation oligomerization.

A key player in mitochondrial respiration, p32, often referred to as C1QBP, is mostly found in the mitochondrial matrix. Previously, we showed that p32 interacts with DLAT in the mitochondria. Here, we found that p32 expression was reduced in ccRCC and suppressed progression and metastasis in ccRCC animal models. We observed that increasing p32 expression led to an increase in oxidative phosphorylation by interacting with DLAT, thus, regulating the activation of the pyruvate dehydrogenase complex (PDHc). Mechanistically, reduced p32 expression, in concert with DLAT, suppresses PDHc activity and the TCA cycle. Furthermore, our research discovered that p32 has a direct binding affinity for copper, facilitating the copper-induced oligomerization of lipo-DLAT specifically in ccRCC cells. This finding reveals an innovative function of the p32/DLAT/copper complex in regulating glycometabolism and the TCA cycle in ccRCC. Importantly, our research provides important new understandings of the underlying molecular processes causing the abnormal mitochondrial metabolism linked to this cancer.

Simple phenylpropanoids: recent advances in biological activities, biosynthetic pathways, and microbial production.

Covering: 2000 to 2023Simple phenylpropanoids are a large group of natural products with primary C6-C3 skeletons. They are not only important biomolecules for plant growth but also crucial chemicals for high-value industries, including fragrances, nutraceuticals, biomaterials, and pharmaceuticals. However, with the growing global demand for simple phenylpropanoids, direct plant extraction or chemical synthesis often struggles to meet current needs in terms of yield, titre, cost, and environmental impact. Benefiting from the rapid development of metabolic engineering and synthetic biology, microbial production of natural products from inexpensive and renewable sources provides a feasible solution for sustainable supply. This review outlines the biological activities of simple phenylpropanoids, compares their biosynthetic pathways in different species (plants, bacteria, and fungi), and summarises key research on the microbial production of simple phenylpropanoids over the last decade, with a focus on engineering strategies that seem to hold most potential for further development. Moreover, constructive solutions to the current challenges and future perspectives for industrial production of phenylpropanoids are presented.

Polysaccharide extracted from cultivated Sanghuangporous vaninii spores using three-phase partitioning with enzyme/ultrasound pretreatment: Physicochemical characteristics and its biological activity in vitro.

Sanghuangporous vaninii, as a valuable dietary supplement and medicinal ingredient, contains abundant bioactive polysaccharides that have health-promoting effects. In the present study, four polysaccharides (SVSPs-C, SVSPs-E, SVSPs-U, and SVSPs-E/U) were extracted for the first time from S. vaninii spores by three-phase partitioning (TPP), enzyme pretreatment before TPP (E-TPP), ultrasonic pretreatment before TPP (U-TPP), and enzyme pretreatment followed by ultrasonic before TPP (E/U-TPP) methods, respectively. Their physicochemical characteristics and in vitro pharmacological functions were determined and compared. Results showed that four TPP-based extraction methods had remarkable impacts on the extraction yield, chemical properties, monosaccharide compositions, and molecular weights (Mw) of SVSPs. Specifically, SVSPs-E/U obtained by E/U-TPP showed the highest extraction yield (25.40 %), carbohydrate content (88.50 %), and the lowest protein content (0.86 %). The four SVSPs had high-Mw (183.8-329.1 kDa) and low-Mw (23.0-156.4 kDa) fractions and mainly consisted of galactose, glucose, and mannose with different contents. In vitro bioactivities assays indicated that SVSPs-E/U possessed stronger antioxidant, hypoglycemic, hypouricemic, immunostimulatory, and antitumor activities than those of SVSPs-C, SVSPs-E, and SVSPs-U. Therefore, our results provide an efficient and promising extraction technique for bioactive polysaccharides from S. vaninii spores, as well as SVSPs had the potential to be applied in functional food, pharmaceutical, and cosmetics fields.

Investigating on sensing mechanism of MoS2-FET biosensors in response to proteins.

Field-effect transistor (FET) biosensors based on two-dimensional materials have gained extensive attention due to their high sensitivity, label-free detection capability, and fast response. Molybdenum disulfide (MoS2), with tunable bandgap, high surface-to-volume ratio, and smooth surface without dangling bonds, is a promising material for FET biosensors. Previous reports have demonstrated the fabrication of MoS2-FET biosensors and their high sensitivity detection of proteins. However, most prior research has focused on the realization of MoS2-FETs for detecting different kinds of proteins or molecules, while comprehensive analysis of the sensing mechanism and dominant device factors of MoS2-FETs in response to proteins is yet to investigate. In this study, we first fabricated MoS2-FET biosensor and detected different types of proteins (immunoglobulin G (IgG),ß-actin, and prostate-specific antigen (PSA)). Secondly, we built the model of the device and analyzed the sensing mechanism of MoS2-FETs in response to proteins. Experimental and modeling results showed that the induced doping effect and gating effect caused by the target protein binding to the device surface were the major influential factors. Specifically, the channel doping concentration and gate voltage (Vg) offset exhibited monotonic change as the concentration of the protein solution increases. For example, the channel doping concentration increased up to ∼37.9% and theVgoffset was ∼-1.3 V with 10-7µgµl-1IgG. The change was less affected by the device size. We also investigated the effects of proteins with opposite acid-base properties (ß-actin and PSA) to IgG on the device sensing mechanism.ß-actin and PSA exhibited behavior opposite to that of IgG. Additionally, we studied the response behavior of MoS2-FETs with different dimensions and dielectric materials (channel length, MoS2thickness, dielectric layer thickness, dielectric layer material) to proteins. The underlying mechanisms were discussed in details. This study provides valuable guidelines for the design and application of MoS2-FET biosensors.

Integrated Proteomics and Single-Cell Mass Cytometry Analysis Dissects the Immune Landscape of Ankylosing Spondylitis.

Mass cytometry is a powerful single-cell technology widely adopted to depict immune cell heterogeneity in different contexts. However, this method is only capable of examining several dozens of proteins simultaneously and requires a prior knowledge of the markers to be analyzed. Here we propose that the integration of mass cytometry with shot-gun proteomics may serve as a valuable tool to achieve an in-depth understanding of the immune system. By implementing such a strategy, we investigated the immune landscape of ankylosing spondylitis (AS), a chronic inflammatory arthritis with unclear etiology. The proteome alteration in peripheral blood mononuclear cells (PBMCs) was investigated by quantitative proteomics, and then mass cytometry analysis was conducted to decipher the immunome by considering the signaling molecules identified with differential expression by proteomics. As a result, we identified a wide spectrum of proteins dysregulated in AS, e.g., upregulation of glycolytic enzymes, downregulation of lipid transporters, and dysregulation of chemokine signaling molecules involved in proinflammatory cytokine production and leucocyte migration. Moreover, the single-cell analysis showed the upregulation of chemokine signaling regulators in subclusters of both innate and adaptive immune cells in AS. In addition, correlation analysis unveiled the interplay among Phenograph-identified subclusters of monocytes, CD4+ T cells, and CD8+ T cells. Taken together, our findings demonstrated that the integration of mass spectrometry-based proteomics and single-cell mass cytometry may serve as a useful tool to reveal clinically relevant information regarding useful targets and cellular phenotypes that could be further exploited to develop novel therapeutic strategies.

Sex Pheromone Receptor-Derived Peptide Biosensor for Efficient Monitoring of the Cotton Bollworm Helicoverpa armigera.

The cotton bollworm, Helicoverpa armigera (H. armigera), causes damage to a wide range of cultivated crops and is one of the pests with the greatest economic importance for global agriculture. Currently, the detection of H. armigera is based on manual sampling. A low limit of detection (LOD), convenient, and real-time monitoring method is urgently needed for its early warning and efficient management. Here, we characterized the amino acid sequence in the sex pheromone receptors (SPRs) recognizing the pheromone components of H. armigera by three-dimensional (3D) modeling and molecular docking. Next, sex pheromone receptor-derived peptides (SPRPs) were synthesized and conjugated to nanotubes by chemical connection. The modified nanotubes were used to fabricate a sensor capable of real-time monitoring of gaseous sex pheromone compounds with a low LOD (∼10 ppb for Z11-16:Ald) and selectivity, and the sensor was able to detect a single live H. armigera. Furthermore, the developed biosensor allowed direct monitoring of the pheromone release dynamics by female H. armigera and showed that the release was instantly reduced in response to light. Here, we report the first demonstration of a biosensing method for detecting gaseous sex pheromones and live H. armigera. The findings show the great potential of the SPRP sensor for broad applications in insect biology study and infestation monitoring.

Profiling of phytohormone-specific microRNAs and characterization of the miR160-ARF1 module involved in glandular trichome development and artemisinin biosynthesis in Artemisia annua.

MicroRNAs (miRNAs) play crucial roles in plant development and secondary metabolism through different modes of sequence-specific interaction with their targets. Artemisinin biosynthesis is extensively regulated by phytohormones. However, the function of phytohormone-responsive miRNAs in artemisinin biosynthesis remains enigmatic. Thus, we combined the analysis of transcriptomics, small RNAs, and the degradome to generate a comprehensive resource for identifying key miRNA-target circuits involved in the phytohormone-induced process of artemisinin biosynthesis in Artemisia annua. In total, 151 conserved and 52 novel miRNAs and their 4132 targets were determined. Based on the differential expression analysis, miR160 was selected as a potential miRNA involved in artemisinin synthesis. Overexpressing MIR160 significantly impaired glandular trichome formation and suppressed artemisinin biosynthesis in A. annua, while repressing its expression resulted in the opposite effect, indicating that miR160 negatively regulates glandular trichome development and artemisinin biosynthesis. RNA ligase-mediated 5' RACE and transient transformation assays showed that miR160 mediates the RNA cleavage of Auxin Response Factor 1 (ARF1) in A. annua. Furthermore, ARF1 was shown to increase artemisinin synthesis by activating AaDBR2 expression. Taken together, our results reveal the intrinsic link between the miR160-ARF1 module and artemisinin biosynthesis, and may expedite the innovation of metabolic engineering approaches for high and stable production of artemisinin in the future.

Genome-wide analysis of AP2/ERF superfamily in Isatis indigotica.

OBJECTIVE: AP2/ERF (APETALA2/ethylene-responsive factor) superfamily is one of the largest gene families in plants and has been reported to participate in various biological processes, such as the regulation of biosynthesis of active lignan. However, few studies have investigated the genome-wide role of the AP2/ERF superfamily in Isatis indigotica. This study establishes a complete picture of the AP2/ERF superfamily in I. indigotica and contributes valuable information for further functional characterization of IiAP2/ERF genes and supports further metabolic engineering. METHODS: To identify the IiAP2/ERF superfamily genes, the AP2/ERF sequences from Arabidopsis thaliana and Brassica rapa were used as query sequences in the basic local alignment search tool. Bioinformatic analyses were conducted to investigate the protein structure, motif composition, chromosome location, phylogenetic relationship, and interaction network of the IiAP2/ERF superfamily genes. The accuracy of omics data was verified by quantitative polymerase chain reaction and heatmap analyses. RESULTS: One hundred and twenty-six putative IiAP2/ERF genes in total were identified from the I. indigotica genome database in this study. By sequence alignment and phylogenetic analysis, the IiAP2/ERF genes were classified into 5 groups including AP2, ERF, DREB (dehydration-responsive element-binding factor), Soloist and RAV (related to abscisic acid insensitive 3/viviparous 1) subfamilies. Among which, 122 members were unevenly distributed across seven chromosomes. Sequence alignment showed that I. indigotica and A. thaliana had 30 pairs of orthologous genes, and we constructed their interaction network. The comprehensive analysis of gene expression pattern in different tissues suggested that these genes may play a significant role in organ growth and development of I. indigotica. Members that may regulate lignan biosynthesis in roots were also preliminarily identified. Ribonucleic acid sequencing analysis revealed that the expression of 76 IiAP2/ERF genes were up- or down-regulated under salt or drought treatment, among which, 33 IiAP2/ERF genes were regulated by both stresses. CONCLUSION: This study undertook a genome-wide characterization of the AP2/ERF superfamily in I. indigotica, providing valuable information for further functional characterization of IiAP2/ERF genes and discovery of genetic targets for metabolic engineering.

Ultrasensitive Flexible Olfactory Receptor-Derived Peptide Sensor for Trimethylamine Detection by the Bending Connection Method.

Trimethylamine (TMA) is a harmful gas that exists ubiquitously in the environment; therefore, the sensitive and specific monitoring of TMA is necessary. In this work, we prepared ultrasensitive flexible sensors for TMA detection based on single-walled carbon nanotubes (SWCNTs) and olfactory receptor-derived peptides (ORPs) on low-cost plastic substrates. A novel bending connection method was developed by intentionally bending the interdigitated electrodes with SWCNTs to form a three-dimensional structure during the ORP-connection process, leading to the exposure of more modification sites. The method showed ∼4.7-fold more effective connection amount of the ORPs to SWCNTs compared to the conventional flat-condition connection method. The flexible ORP-SWCNT sensors could significantly improve the limit of detection for gaseous TMA from the reported lowest limit of 10 parts per quadrillion (ppq) to 0.1 ppq. The flexible ORP sensors also exhibited excellent sensitivity to vaporized TMA standards and TMA generated by different kinds of foods under different bending conditions. The results showed that the bending connection method in this work was effective for ultrasensitive flexible ORP sensors and their associated applications.

Biosynthesis and regulation of diterpenoids in medicinal plants.

Plant diterpenoids are widely distributed and abundant natural products with diverse structures and functions in nature, which have been commonly used in pharmaceutical, agricultural and industrial production. In recent years, plant diterpenoids have attracted increasing attention, including their biosynthetic pathways, transcriptional regulatory networks, and biological functions. Herein, the biosynthetic pathways of diterpenoids are summarized in a modular fashion. Further, the regulatory network between diterpene biosynthesis and environmental factors is reviewed. Insights into diterpene metabolism may drive elucidation of complex active diterpene pathways and serve as a knowledge repository for metabolic engineering and cell factory construction.

Hydrophilic glutathione-modified flower-like hollow covalent organic frameworks for highly efficient capture of N-linked glycopeptides.

Highly efficient enrichment of N-glycopeptides from complicated biosamples based on mass spectrometry is essential for biomedical applications, especially in disease biomarker research. In this work, glutathione (GSH)-modified hierarchical flower-like hollow covalent organic frameworks loaded with Au nanoparticles (HFH-COFs@Au@GSH) were synthesized for N-glycopeptide enrichment. Due to the abundant accessibility sites, high specific surface area, and inherent high stability of the hierarchical flower-like hollow structure, a large number of Au NPs and hydrophilic GSH can be modified on the HFH-COFs. The HFH-COFs@Au@GSH displayed excellent hydrophilicity and remarkable enrichment performance for N-glycopeptides: low detection limit (0.1 fmol µL-1), large adsorption capacity (200 µg mg-1), great selectivity (1 : 1000, HRP to BSA), and good reusability (at least 5 times). Furthermore, the HFH-COFs@Au@GSH were successfully applied to capture N-linked glycopeptides in human serum, and 308 N-glycosylation peptides corresponding to 84 N-glycosylation proteins with 123 N-glycosylation sites were detected. Gene ontology analyses were used to elucidate the cellular component, biological process and molecular function of detected glycoproteins in human serum, demonstrating the great potential of the HFH-COFs@Au@GSH in N-glycopeptide enrichment for glycoproteomic analysis of complex biological samples.

Quantitative proteomic analysis of oxaliplatin induced peripheral neurotoxicity.

Oxaliplatin (OXA)-induced peripheral neurotoxicity (OIPN) is a high-incidence and dose-dependent adverse reaction during OXA treatment. Its underlying mechanisms remain unclear, and no effective treatment or prevention therapies are currently available. Here, we employed a data independent acquisition (DIA)-based quantitative proteomic strategy to investigate the global proteome alterations in the dorsal root ganglion (DRG) tissues from mice injected with OXA for different periods. We identified 1128 differentially regulated proteins that were divided into six subclusters according to their alteration trends. Interestingly, these proteins were involved in cellular processes such as cell cycle, ribosomal stress, metabolism, and ion transport. In addition, OXA administration induced abundance changes of ion channels and proteins associated with mitochondrial function and reactive oxygen species production. Furthermore, we investigated the effects of diroximel fumarate (DRF), an FDA-approved oral fumarate drug for the treatment of relapsing forms of multiple sclerosis. Our findings showed that DRF could effectively ameliorate symptoms of OIPN and reduce the level of oxidative stress in mice. Taken together, our study systematically mapped the proteome alteration associated with the neural toxicity of OXA, and the findings could be leveraged to better understand the mechanisms of OIPN and to develop more effect treatment therapies. SIGNIFICANCE: Oxaliplatin (OXA)-induced peripheral neurotoxicity (OIPN) is a high-incidence and dose-dependent adverse reaction with unclear mechanism. Here we employed a data independent acquisition (DIA)-based quantitative proteomic strategy to explore the proteome changes in dorsal root ganglion (DRG) tissues from mice treated by OXA. The findings provided novel insights regarding the mechanisms of OIPN. For example, our data showed that OXA induced a broad disturbance in metabolism, particularly in glycolysis and amino acid metabolism. Additionally, we observed abundance changes of many ion channels and proteins associated with mitochondrial function and reactive oxygen species production. Furthermore, this study provided the first evidence for the possibility of repositioning diroximel fumarate (DRF) for treating OIPN.

C1QBP regulates apoptosis of renal cell carcinoma via modulating xanthine dehydrogenase (XDH) mediated ROS generation.

Background: Complement component 1 Q subcomponent binding protein (C1QBP) plays a vital role in the progression and metabolism of cancer. Studies have shown that xanthine dehydrogenase (XDH)-derived reactive oxygen species (ROS) accelerates tumor growth, and also induces mutations or produces cytotoxic effects concurrently. However, the role of C1QBP in metabolism, oxidative stress, and apoptosis of renal cell carcinoma (RCC) cells have not yet been explored. Methods: Metabolomics assay was applied to investigate the role of C1QBP in RCC metabolism. C1QBP knockdown and overexpression cells were established via lentiviral infection and subjected to apoptosis and ROS assay in vitro. RNA stability assay was applied to characterize the mechanism of C1QBP regulating XDH transcription. In vivo, orthotopic tumor xenografts assay was performed to investigate the role of C1QBP in RCC progression. Results: Metabolomics investigation revealed that C1QBP dramatically diminished the hypoxanthine content in RCC cells. C1QBP promoted the mRNA and protein expression of hypoxanthine catabolic enzyme XDH. Meanwhile, C1QBP may affect XDH transcription by regulating the mRNA level of XDH transcriptional stimulators IL-6, TNF-α, and IFN-γ. Moreover, the expression of C1QBP and XDH was lower in RCC tumors compared with the tumor-associated normal tissues, and their down-regulation was associated with higher Fuhrman grade. C1QBP significantly increased ROS level, apoptosis, and the expression of apoptotic proteins such as cleaved caspase-3 and bax/bcl2 via regulating XDH. Conclusion: C1QBP promotes the catabolism of hypoxanthine and elevates the apoptosis of RCC cells by modulating XDH-mediated ROS generation.

In-depth Profiling and Quantification of the Lysine Acetylome in Hepatocellular Carcinoma with a Trapped Ion Mobility Mass Spectrometer.

Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related death worldwide with limited therapeutic options. Comprehensive investigation of protein posttranslational modifications in HCC is still limited. Lysine acetylation is one of the most common types of posttranslational modification involved in many cellular processes and plays crucial roles in the regulation of cancer. In this study, we analyzed the proteome and K-acetylome in eight pairs of HCC tumors and normal adjacent tissues using a timsTOF Pro instrument. As a result, we identified 9219 K-acetylation sites in 2625 proteins, of which 1003 sites exhibited differential acetylation levels between tumors and normal adjacent tissues. Interestingly, many novel tumor-specific K-acetylation sites were characterized, for example, filamin A (K865), filamin B (K697), and cofilin (K19), suggesting altered activities of these cytoskeleton-modulating molecules, which may contribute to tumor metastasis. In addition, we observed an overall suppression of protein K-acetylation in HCC tumors, especially for enzymes from various metabolic pathways, for example, glycolysis, tricarboxylic acid cycle, and fatty acid metabolism. Moreover, the expression of deacetylase sirtuin 2 (SIRT2) was upregulated in HCC tumors, and its role of deacetylation in HCC cells was further explored by examining the impact of SIRT2 overexpression on the proteome and K-acetylome in Huh7 HCC cells. SIRT2 overexpression reduced K-acetylation of proteins involved in a wide range of cellular processes, including energy metabolism. Furthermore, cellular assays showed that overexpression of SIRT2 in HCC cells inhibited both glycolysis and oxidative phosphorylation. Taken together, our findings provide valuable information to better understand the roles of K-acetylation in HCC and to treat this disease by correcting the aberrant acetylation patterns.

A Single-Cell Atlas of Tumor-Infiltrating Immune Cells in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with limited treatment options. To guide the design of more effective immunotherapy strategies, mass cytometry was employed to characterize the cellular composition of the PDAC-infiltrating immune cells. The expression of 33 protein markers was examined at the single-cell level in more than two million immune cells from four types of clinical samples, including PDAC tumors, normal pancreatic tissues, chronic pancreatitis tissues, and peripheral blood. Based on the analyses, we identified 23 distinct T-cell phenotypes, with some cell clusters exhibiting aberrant frequencies in the tumors. Programmed cell death protein 1 (PD-1) was extensively expressed in CD4+ and CD8+ T cells and coexpressed with both stimulatory and inhibitory immune markers. In addition, we observed elevated levels of functional markers, such as CD137L and CD69, in PDAC-infiltrating immune cells. Moreover, the combination of PD-1 and CD8 was used to stratify PDAC tumors from The Cancer Genome Atlas database into three immune subtypes, with S1 (PD-1+CD8+) exhibiting the best prognosis. Further analysis suggested distinct molecular mechanisms for immune exclusion in different subtypes. Taken together, the single-cell protein expression data depicted a detailed cell atlas of the PDAC-infiltrating immune cells and revealed clinically relevant information regarding useful cell phenotypes and targets for immunotherapy development.

Polystyrene nanoplastics induce profound metabolic shift in human cells as revealed by integrated proteomic and metabolomic analysis.

Nanoplastics (NPLs) are widespread in our environment. However, their impacts on human health and precise toxicity mechanisms remain poorly understood. Here we studied the internalization, release, and cytotoxicity of polystyrene nanoplastics (PSNPs) using the renal tubular epithelial cell line HKC and human derived liver cell line HL-7702. We also employed an integrated proteomic and metabolomic approach to investigate the potential biological effects of PSNPs on HKC cells. The abundances of 4770 proteins and 100 metabolites were quantified, with 785 proteins and 17 metabolites detected with altered levels in response to PSNPs. Most of the differential proteins and metabolites were enriched in a variety of metabolic pathways, for example, glycolysis, citrate cycle, oxidative phosphorylation, and amino acid metabolism, suggesting the potential effects of NPLs on global cellular metabolism shift in human cells. The altered energy metabolism induced by PSNPs was further confirmed by a Seahorse analysis. Moreover, lysosomal distribution study and western blotting showed that mTORC1 signaling, a central regulator of cellular metabolism, was inhibited upon nanoplastic exposure, likely serving as the link between lysosome dysfunction and metabolic defects. Taken together, our findings systematically mapped the key molecular changes induced by PSNPs in human cells and provide comprehensive biological insights for the risk estimation of NPLs contamination.