Integrative Profiling Research Articles

Integrative profiling of metabolome and transcriptome of skeletal muscle after acute exercise intervention in mice.

This study aims to explore the molecular regulatory mechanisms of acute exercise in the skeletal muscle of mice. Male C57BL/6 mice were randomly assigned to the control group, and the exercise group, which were sacrificed immediately after an acute bout of exercise. The study was conducted to investigate the metabolic and transcriptional profiling in the quadriceps muscles of mice. The results demonstrated the identification of 34 differentially expressed metabolites (DEMs), with 28 upregulated and 6 downregulated, between the two groups. Metabolic pathway analysis revealed that these DEMs were primarily enriched in several, including the citrate cycle, propanoate metabolism, and lysine degradation pathways. In addition, the results showed a total of 245 differentially expressed genes (DEGs), with 155 genes upregulated and 90 genes downregulated. KEGG analysis indicated that these DEGs were mainly enriched in various pathways such as ubiquitin mediated proteolysis and FoxO signaling pathway. Furthermore, the analysis revealed significant enrichment of DEMs and DEGs in signaling pathways such as protein digestion and absorption, ferroptosis signaling pathway. In summary, the identified multiple metabolic pathways and signaling pathways were involved in the exercise-induced physiological regulation of skeletal muscle, such as the TCA cycle, oxidative phosphorylation, protein digestion and absorption, the FoxO signaling pathway, ubiquitin mediated proteolysis, ferroptosis signaling pathway, and the upregulation of KLF-15, FoxO1, MAFbx, and MuRF1 expression could play a critical role in enhancing skeletal muscle proteolysis.

336.6: Integrative omic profiling and analyses in two pig heart to human xenotransplants

336.6: Integrative omic profiling and analyses in two pig heart to human xenotransplants

Multimodal analysis of cell-free DNA whole-methylome sequencing for cancer detection and localization

Multimodal epigenetic characterization of cell-free DNA (cfDNA) could improve the performance of blood-based early cancer detection. However, integrative profiling of cfDNA methylome and fragmentome has been technologically challenging. Here, we adapt an enzyme-mediated methylation sequencing method for comprehensive analysis of genome-wide cfDNA methylation, fragmentation, and copy number alteration (CNA) characteristics for enhanced cancer detection. We apply this method to plasma samples of 497 healthy controls and 780 patients of seven cancer types and develop an ensemble classifier by incorporating methylation, fragmentation, and CNA features. In the test cohort, our approach achieves an area under the curve value of 0.966 for overall cancer detection. Detection sensitivity for early-stage patients achieves 73% at 99% specificity. Finally, we demonstrate the feasibility to accurately localize the origin of cancer signals with combined methylation and fragmentation profiling of tissue-specific accessible chromatin regions. Overall, this proof-of-concept study provides a technical platform to utilize multimodal cfDNA features for improved cancer detection.

DARESOME enables concurrent profiling of multiple DNA modifications with restriction enzymes in single cells and cell-free DNA.

5-Methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) are the most abundant DNA modifications that have important roles in gene regulation. Detailed studies of these different epigenetic marks aimed at understanding their combined effects and dynamic interconversion are, however, hampered by the inability of current methods to simultaneously measure both modifications, particularly in samples with limited quantities. We present DNA analysis by restriction enzyme for simultaneous detection of multiple epigenomic states (DARESOME), an assay based on modification-sensitive restriction digest and sequential tag ligation that can concurrently perform quantitative profiling of unmodified cytosine, 5mC, and 5hmC in CCGG sites genome-wide. DARESOME reveals the opposing roles of 5mC and 5hmC in gene expression regulation as well as their interconversion during aging in mouse brain. Implementation of DARESOME in single cells demonstrates pronounced 5hmC strand bias that reflects the semiconservative replication of DNA. Last, we showed that DARESOME enables integrative genomic, 5mC, and 5hmC profiling of cell-free DNA that uncovered multiomics cancer signatures in liquid biopsy.

The importance of escalating molecular diagnostics in patients with low-grade pediatric brain cancer.

Pilocytic astrocytomas are the most common pediatric brain tumors, typically presenting as low-grade neoplasms. We report two cases of pilocytic astrocytoma with atypical tumor progression. Case 1 involves a 12-yr-old boy with an unresectable suprasellar tumor, negative for BRAF rearrangement but harboring a BRAF p.V600E mutation. He experienced tumor size reduction and stable disease following dabrafenib treatment. Case 2 describes a 6-yr-old boy with a thalamic tumor that underwent multiple resections, with no actionable driver detected using targeted next-generation sequencing. Whole-genome and RNA-seq analysis identified an internal tandem duplication in FGFR1 and RAS pathway activation. Future management options include FGFR1 inhibitors. These cases demonstrate the importance of escalating molecular diagnostics for pediatric brain cancer, advocating for early reflexing to integrative whole-genome sequencing and transcriptomic profiling when targeted panels are uninformative. Identifying molecular drivers can significantly impact treatment decisions and improve patient outcomes.

Integrative HPLC profiling and transcriptome analysis revealed insights into anthocyanin accumulation and key genes at three developmental stages of black rice (Oryza sativa. L) caryopsis.

Anthocyanins are plants' secondary metabolites belonging to the flavonoid class with potential health-promoting properties. They are greatly employed in the food industry as natural alternative food colorants for dairy and ready-to-eat desserts and pH indicators. These tremendous advantages make them economically important with increasing market trends. Black rice is a rich source of anthocyanin that can be used to ensure food and nutritional security around the world. However, research on anthocyanin accumulation and gene expression during rice caryopsis development is lacking. In this study, we combined high-performance liquid chromatography (HPLC) and transcriptome analysis to profile the changes in anthocyanin content and gene expression dynamics at three developmental stages (milky, doughy, and mature). Our results showed that anthocyanin accumulation started to be visible seven days after flowering (DAF), increased rapidly from milky (11 DAF) to dough stage, then started decreasing after the peak was attained at 18 DAF. RNA-seq showed that 519 out of 14889, 477 out of 17914, and 1614 out of 18810 genes were uniquely expressed in the milky, doughy, and mature stages, respectively. We performed three pairwise comparisons: milky vs. dough, milky vs. mature, and dough vs. mature, and identified 6753, 9540, and 2531 DEGs, respectively. The DEGs' abundance was higher in milky vs. mature, with 5527 up-regulated genes and 4013 down-regulated genes, while it was smaller in dough vs. mature, with 1419 up-regulated genes and 1112 down-regulated DEGs. This result was consistent with the changes in anthocyanin profiling, and the expression of structural, regulatory, and transporter genes involved in anthocyanin biosynthesis showed their highest expression at the dough stage. Through the gene expression profile and protein interaction network, we deciphered six main contributors of the anthocyanin peak observed at dough stage, including OsANS, OsDFR, OsGSTU34, OsMYB3, OsbHLH015, and OsWD40-50. This study is the first to report the investigation of anthocyanin and gene expression at three developmental stages of black rice caryopsis. The findings of this study could aid in predicting the best harvesting time to achieve maximum anthocyanin content and the best time to collect samples for various gene expression analysis, laying the groundwork for future research into the molecular mechanisms underlying rice caryopsis coloration.

Integrative transcriptomic profiling of mRNA, miRNA, circRNA, and lncRNA in alveolar macrophages isolated from PRRSV-infected porcine.

The porcine reproductive and respiratory syndrome virus (PRRSV) continues to pose a significant threat to the global swine industry, attributed largely to its immunosuppressive properties and the chronic nature of its infection. The absence of effective vaccines and therapeutics amplifies the urgency to deepen our comprehension of PRRSV's intricate pathogenic mechanisms. Previous transcriptomic studies, although informative, are partially constrained by their predominant reliance on in vitro models or lack of long-term infections. Moreover, the role of circular RNAs (circRNAs) during PRRSV invasion is yet to be elucidated. In this study, we employed an in vivo approach, exposing piglets to a PRRSV challenge over varied durations of 3, 7, or 21 days. Subsequently, porcine alveolar macrophages were isolated for a comprehensive transcriptomic investigation, examining the expression patterns of mRNAs, miRNAs, circRNAs, and long non-coding RNAs (lncRNAs). Differentially expressed RNAs from all four categories were identified, underscoring the dynamic interplay among these RNA species during PRRSV infection. Functional enrichment analyses indicate that these differentially expressed RNAs, as well as their target genes, play a pivotal role in immune related pathways. For the first time, we integrated circRNAs into the lncRNA-miRNA-mRNA relationship, constructing a competitive endogenous RNA (ceRNA) network. Our findings highlight the immune-related genes, CTLA4 and SAMHD1, as well as their associated miRNAs, lncRNAs, and circRNAs, suggesting potential therapeutic targets for PRRS. Importantly, we corroborated the expression patterns of selected RNAs through RT-qPCR, ensuring consistency with our transcriptomic sequencing data. This study sheds lights on the intricate RNA interplay during PRRSV infection and provides a solid foundation for future therapeutic strategizing.

Integrative multi-omic cancer profiling reveals DNA methylation patterns associated with therapeutic vulnerability and cell-of-origin

DNA methylation plays a critical role in establishing and maintaining cellular identity. However, it is frequently dysregulated during tumor development and is closely intertwined with other genetic alterations. Here, we leveraged multi-omic profiling of 687 tumors and matched non-involved adjacent tissues from the kidney, brain, pancreas, lung, head and neck, and endometrium to identify aberrant methylation associated with RNA and protein abundance changes and build a Pan-Cancer catalog. We uncovered lineage-specific epigenetic drivers including hypomethylated FGFR2 in endometrial cancer. We showed that hypermethylated STAT5A is associated with pervasive regulon downregulation and immune cell depletion, suggesting that epigenetic regulation of STAT5A expression constitutes a molecular switch for immunosuppression in squamous tumors. We further demonstrated that methylation subtype-enrichment information can explain cell-of-origin, intra-tumor heterogeneity, and tumor phenotypes. Overall, we identified cis-acting DNA methylation events that drive transcriptional and translational changes, shedding light on the tumor's epigenetic landscape and the role of its cell-of-origin.

Maternal voluntary wheel running modulates glucose homeostasis, the gut microbiota and its derived fecal metabolites in offspring.

Maternal overnutrition can dramatically increase the susceptibility of offspring to metabolic diseases, whereas maternal exercise may improve glucose metabolism in offspring. However, the underlying mechanism programming the intergenerational effects of maternal exercise on the benefits of glucose metabolism has not been fully elaborated. C57BL/6 female mice were randomly assigned to four subgroups according to a diet and exercise paradigm before and during pregnancy as follows: NC (fed with normal chow diet and sedentary), NCEx (fed with normal chow diet and running), HF (fed with high-fat diet and sedentary), and HFEx (fed with high-fat diet and running). Integrative 16S rDNA sequencing and mass spectrometry-based metabolite profiling were synchronously performed to characterize the effects of maternal exercise on the gut microbiota composition and metabolite alterations in offspring. Maternal exercise, acting as a natural pharmaceutical intervention, prevented deleterious effects on glucose metabolism in offspring. 16S rDNA sequencing revealed remarkable changes in the gut microbiota composition in offspring. Metabolic profiling indicated multiple altered metabolites, which were enriched in butanoate metabolism signaling in offspring. We further found that maternal exercise could mediate gene expression related to intestinal gluconeogenesis in offspring. In conclusion, our study indicated that maternal running significantly improved glucose metabolism in offspring and counteracted the detrimental effects of maternal high-fat feeding before and during pregnancy. We further demonstrated that maternal voluntary wheel running could integratively program the gut microbiota composition and fecal metabolite changes and then regulate butanoate metabolism and mediate intestinal gluconeogenesis in offspring.

Integrative Profiling of Lysine Acylome in Sepsis-Induced Liver Injury.

Sepsis is one of the life-threatening diseases worldwide. Despite the continuous progress in medicine, the specific mechanism of sepsis remains unclear. A key strategy of pathogens is to use post-translational modification to modulate host factors critical for infection. We employed global immunoprecipitation technology for lysine acetylation (Kac), succinylation (Ksu), and malonylation (Kmal) for the first global lysine acylation (Kacy) analysis in a cecum ligation and puncture (CLP) model in mouse. This was performed to reveal the pathogenic mechanism of integrative Kacy and the changes in modification sites. In total, 2230 sites of 1,235 Kac proteins, 1,887 sites of 433 Ksu proteins, and 499 sites of 276 Kmal proteins were quantified and normalized by their protein levels. We focused on 379 sites in 219 upregulated proteins as the integrative Kacy sites of Kac, Ksu, and Kmal on the basis of sirtuins decreased in the CLP group. KEGG pathway analysis of integrative Kacy in 219 upregulated proteins revealed three central metabolic pathways: glycolysis/gluconeogenesis, pyruvate metabolism, and tricarboxylic acid cycle. These findings reveal the key pathogenic mechanism of integrative PTM alteration in terms of the decreased sirtuins level and provide an important foundation for an in-depth study of the biological function of Kacy in sepsis.

Integrative Transcriptomic Profiling of the Wilms Tumor.

Our study aimed to identify relevant transcriptomic biomarkers for the Wilms tumor, the most common pediatric kidney cancer, independent of the histological type and stage. Using next-generation sequencing, we analyzed the miRNA profiles of 74 kidney samples, which were divided into two independent groups: fresh frozen tissue and formalin-fixed paraffin-embedded tissue samples. Subsequent mRNA expression profiling and pathway analysis were performed to establish the interplay and potential involvement of miRNAs and mRNA in the Wilms tumor. Comparative analysis, irrespective of post-dissection tissue processing, revealed 41 differentially expressed miRNAs, with 27 miRNAs having decreased expression and 14 miRNAs having increased expression in the Wilms tumor tissue compared to healthy kidney tissue. Among global mRNA transcriptomic profile differences, cross-sectional analysis suggested a limited list of genes potentially regulated by differentially expressed miRNAs in the Wilms tumor. This study identified the comprehensive miRNA and mRNA profile of the Wilms tumor using next-generation sequencing and bioinformatics approach, providing better insights into the pathogenesis of the Wilms tumor. The identified Wilms tumor miRNAs have potential as biomarkers for the diagnosis and treatment of the Wilms tumor, regardless of histological subtype and disease stage.

Integrative Metabolic and Transcriptomic Profiling in Camellia oleifera and Camellia meiocarpa Uncover Potential Mechanisms That Govern Triacylglycerol Degradation during Seed Desiccation.

Camellia seed oil is a top-end quality of cooking oil in China. The oil quality and quantity are formed during seed maturation and desiccation. So far, it remains largely unresolved whether lipid degradation occurs and contributes to Camellia oil traits. In this study, three different Camellia germplasms, C. oleifera cv. Min 43 (M43), C. meiocarpa var. Qingguo (QG), and C. meiocarpa cv Hongguo (HG) were selected, their seed oil contents and compositions were quantified across different stages of seed desiccation. We found that at the late stage of desiccation, M43 and QG lost a significant portion of seed oil, while such an event was not observed in HG. To explore the molecular bases for the oil loss In M43, the transcriptomic profiling of M43 and HG was performed at the early and the late seed desiccation, respectively, and differentially expressed genes (DEGs) from the lipid metabolic pathway were identified and analyzed. Our data demonstrated that different Camellia species have diverse mechanisms to regulate seed oil accumulation and degradation, and that triacylglycerol-to-terpenoid conversion could account for the oil loss in M43 during late seed desiccation.

Integrative profiling of extrachromosomal circular DNA in placenta and maternal plasma provides insights into the biology of fetal growth restriction and reveals potential biomarkers.

Introduction: Fetal growth restriction (FGR) is a placenta-mediated pregnancy complication that predisposes fetuses to perinatal complications. Maternal plasma cell-free DNA harbors DNA originating from placental trophoblasts, which is promising for the prenatal diagnosis and prediction of pregnancy complications. Extrachromosomal circular DNA (eccDNA) is emerging as an ideal biomarker and target for several diseases. Methods: We utilized eccDNA sequencing and bioinformatic pipeline to investigate the characteristics and associations of eccDNA in placenta and maternal plasma, the role of placental eccDNA in the pathogenesis of FGR, and potential plasma eccDNA biomarkers of FGR. Results: Using our bioinformatics pipelines, we identified multi-chromosomal-fragment and single-fragment eccDNA in placenta, but almost exclusively single-fragment eccDNA in maternal plasma. Relative to that in plasma, eccDNA in placenta was larger and substantially more abundant in exons, untranslated regions, promoters, repetitive elements [short interspersed nuclear elements (SINEs)/Alu, SINEs/mammalian-wide interspersed repeats, long terminal repeats/endogenous retrovirus-like elements, and single recognition particle RNA], and transcription factor binding motifs. Placental multi-chromosomal-fragment eccDNA was enriched in confident enhancer regions predicted to pertain to genes in apoptosis, energy, cell growth, and autophagy pathways. Placental eccDNA-associated genes whose abundance differed between the FGR and control groups were associated with immunity-related gene ontology (GO) terms. The combined analysis of plasma and placental eccDNA-associated genes in the FGR and control groups led to the identification of potential biomarkers that were assigned to the GO terms of the epigenetic regulation of gene expression and nutrient-related processes, respectively. Conclusion: Together, our results highlight links between placenta functions and multi-chromosomal-fragment and single-fragment eccDNA. The integrative analysis of placental and plasma eccDNA confirmed the potential of these molecules as disease-specific biomarkers of FGR.

STAT3-CD163 cross-talk exhibits promising biomarkers for a progressive ischemic cardiomyopathy: integrative computational and gene expression profiling based on Gene Expression Omnibus datasets

Background and purpose: Ischemic heart disease frequently leads to heart failure, often resulting in death. In this study, we aimed to identify common hub mRNAs and pathways involved in the pathological progression of ischemic cardiomyopathy (ICM). Methods: Validation quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was carried out on peripheral blood and left ventricle specimens from patients in three groups with stable coronary artery disease (CAD), myocardial infarction (MI), and ICM and compared with corresponding controls. qRT-PCR was preceded by computational analysis of eight high-throughput RNA sequencing and microarray datasets from 499 patients and 233 controls, to determine possible common biologically meaningful differentially expressed genes (DEGs). To determine the potential pathological pathways, we performed Gene Ontology functional annotation, pathway enrichment analysis, protein–protein interaction (PPI) analysis, and constructed transcriptional factor/miRNA regulatory networks. Finally, approved drugs were screened. Results: Fifteen common DEGs with P < 0.01 were identified. STAT3, CEBPD, GLUL, and CD163 were hub-enriched mRNAs with an interaction score ≥ 0.50. Our qRT-PCR analysis showed an increased expression of STAT3 in all three patient groups and CD163, mainly in cardiac samples, in a remarkably ascending manner. Interaction modules showed co-regulators supporting high STAT3-CD163 connectivity, suggesting a potential role for STAT3-CD163 cross-talk-mediated inflammatory responses in ICM progression. Conclusions: Our results provided a novel perspective for understanding the underlying mechanisms of ICM progression and exploring new therapeutic agents. Clinical trial registration: URL: www.clinicaltrials.gov. Unique identifier: NCT05508269.

Chemoproteomics and Phosphoproteomics Profiling Reveals Salvianolic Acid A as a Covalent Inhibitor of mTORC1.

Salvianolic acid A (SAA), a major active ingredient of Salvia miltiorrhiza Bunge (Danshen), displays strong antiproliferative activity against cancer cells. However, their protein targets remain unknown. Here, we deconvoluted the protein targets of SAA using chemoproteomics and phosphoproteomics. By using alkynylated SAA as a probe, we discovered that SAA is a covalent ligand that can modify cellular proteins via its electrophilic α,β-unsaturated ester moiety. The subsequent chemoproteomics profiling revealed that 46 proteins were covalently modified by SAA, including Raptor, a subunit of mTORC1 for recruiting substrates for mTORC1. Although gene ontology enrichment analysis of these proteins suggested that SAA displays a promiscuous protein interaction, phosphoproteomics profiling revealed that the SAA modulated phosphoproteins were mainly enriched in the signaling pathways of PI3K-Akt-mTOR, which is closely related to cell growth and proliferation. This was confirmed by the biochemical assay with purified mTORC1, a Western blot assay with phospho-specific antibodies, and a cellular thermal shift assay. Our work discovered that SAA is a covalent ligand for protein modification and mTORC1 is one of its targets. Moreover, our work demonstrated that the integrative profiling of chemoproteomics and phosphoproteomics can be a powerful tool for target deconvolution for bioactive natural products.

Integrative profiling analysis reveals prognostic significance, molecular characteristics, and tumor immunity of angiogenesis-related genes in soft tissue sarcoma.

Soft tissue sarcoma (STS) is a class of malignant tumors originating from mesenchymal stroma with a poor prognosis. Accumulating evidence has proved that angiogenesis is an essential hallmark of tumors. Nevertheless, there is a paucity of comprehensive research exploring the association of angiogenesis-related genes (ARGs) with STS. The ARGs were extracted from previous literature, and the differentially expressed ARGs were screened for subsequent analysis. Next, the least absolute shrinkage and selection operator (LASSO) and Cox regression analyses were conducted to establish the angiogenesis-related signature (ARSig). The predictive performance of the novel ARSig was confirmed using internal and external validation, subgroup survival, and independent analysis. Additionally, the association of the ARSig with the tumor immune microenvironment, tumor mutational burden (TMB), and therapeutic response in STS were further investigated. Notably, we finally conducted in vitro experiments to verify the findings from the bioinformatics analysis. A novel ARSig is successfully constructed and validated. The STS with a lower ARSig risk score in the training cohort has an improved prognosis. Also, consistent results were observed in the internal and external cohorts. The receiver operating characteristic (ROC) curve, subgroup survival, and independent analysis further indicate that the novel ARSig is a promising independent prognostic predictor for STS. Furthermore, it is proved that the novel ARSig is relevant to the immune landscape, TMB, immunotherapy, and chemotherapy sensitivity in STS. Encouragingly, we also validate that the signature ARGs are significantly dysregulated in STS, and ARDB2 and SRPK1 are closely connected with the malignant progress of STS cells. In sum, we construct a novel ARSig for STS, which could act as a promising prognostic factor for STS and give a strategy for future clinical decisions, immune landscape, and personalized treatment of STS.

Integrative profiling of gene expression and chromatin accessibility elucidates specific transcriptional networks in porcine neutrophils.

Neutrophils are vital components of the immune system for limiting the invasion and proliferation of pathogens in the body. Surprisingly, the functional annotation of porcine neutrophils is still limited. The transcriptomic and epigenetic assessment of porcine neutrophils from healthy pigs was performed by bulk RNA sequencing and transposase accessible chromatin sequencing (ATAC-seq). First, we sequenced and compared the transcriptome of porcine neutrophils with eight other immune cell transcriptomes to identify a neutrophil-enriched gene list within a detected neutrophil co-expression module. Second, we used ATAC-seq analysis to report for the first time the genome-wide chromatin accessible regions of porcine neutrophils. A combined analysis using both transcriptomic and chromatin accessibility data further defined the neutrophil co-expression network controlled by transcription factors likely important for neutrophil lineage commitment and function. We identified chromatin accessible regions around promoters of neutrophil-specific genes that were predicted to be bound by neutrophil-specific transcription factors. Additionally, published DNA methylation data from porcine immune cells including neutrophils were used to link low DNA methylation patterns to accessible chromatin regions and genes with highly enriched expression in porcine neutrophils. In summary, our data provides the first integrative analysis of the accessible chromatin regions and transcriptional status of porcine neutrophils, contributing to the Functional Annotation of Animal Genomes (FAANG) project, and demonstrates the utility of chromatin accessible regions to identify and enrich our understanding of transcriptional networks in a cell type such as neutrophils.

Metabolome and transcriptome analyses reveal the molecular mechanisms of LcMYB1 regulating anthocyanin accumulation in litchi hairy roots

Agrobacterium rhizogenes-mediated hairy root culture offer a promising approach for gene function analysis and production of plant secondary metabolites. Here, we obtained red litchi hairy roots using A. rhizogenes-mediated LcMYB1 transformation. Using high performance liquid chromatography, the main anthocyanins in the red hairy roots were determined to be cyanidin 3-rutinoside and cyanidin 3-glucoside. A total of 164 metabolites were significantly upregulated or downregulated in the red hairy roots, which were mostly involved in flavone and flavonol pathway, and flavonoid pathway. The transcriptome analysis revealed 472 differentially expressed genes (DEGs). Up-regulated genes were considerably enriched in anthocyanin, flavone and flavonol biosynthesis. Integrative metabolite profiling and transcriptome analyses showed that LcF3′H, LcUFGT1, and LcGST4 were key structural genes in anthocyanin biosynthesis. However, the expression of Cinnamyl-alcohol dehydrogenase (CAD) and Peroxidase (POD) leading to the production of lignin were significantly down-regulated, suggesting flavonoids and lignin compete with each other in the phenylpropanoid pathway. A total of 52 DEGs were identified as transcription factors. Correlation analysis showed that 8 transcription factors were positively correlated with LcUFGT1, and LcGST4, involving in anthocyanin biosynthesis. These findings clarify the molecular mechanisms of LcMYB1 regulating anthocyanin accumulation in litchi hairy roots.

Integrative proteomic profiling of lung tissues and blood in acute respiratory distress syndrome.

Acute respiratory distress syndrome and acute lung injury (ARDS/ALI) still lack a recognized diagnostic test and pharmacologic treatments that target the underlying pathology. To explore the sensitive non-invasive biomarkers associated with pathological changes in the lung of direct ARDS/ALI, we performed an integrative proteomic analysis of lung and blood samples from lipopolysaccharide (LPS)-induced ARDS mice and COVID-19-related ARDS patients. The common differentially expressed proteins (DEPs) were identified based on combined proteomic analysis of serum and lung samples in direct ARDS mice model. The clinical value of the common DEPs was validated in lung and plasma proteomics in cases of COVID-19-related ARDS. We identified 368 DEPs in serum and 504 in lung samples from LPS-induced ARDS mice. Gene ontology (GO) classification and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that these DEPs in lung tissues were primarily enriched in pathways, including IL-17 and B cell receptor signaling pathways, and the response to stimuli. In contrast, DEPs in the serum were mostly involved in metabolic pathways and cellular processes. Through network analysis of protein-protein interactions (PPI), we identified diverse clusters of DEPs in the lung and serum samples. We further identified 50 commonly upregulated and 10 commonly downregulated DEPs in the lung and serum samples. Internal validation with a parallel-reacted monitor (PRM) and external validation in the Gene Expression Omnibus (GEO) datasets further showed these confirmed DEPs. We then validated these proteins in the proteomics of patients with ARDS and identified six proteins (HP, LTA4H, S100A9, SAA1, SAA2, and SERPINA3) with good clinical diagnostic and prognostic value. These proteins can be viewed as sensitive and non-invasive biomarkers associated with lung pathological changes in the blood and could potentially serve as targets for the early detection and treatment of direct ARDS especially in hyperinflammatory subphenotype.

Deciphering Cell-Cell Interactions with Integrative Single-Cell Secretion Profiling.

Cell-cell interactions are the fundamental behaviors to regulate cellular activities. A comprehensive evaluation of intercellular interactions requires direct profiling of various signaling behaviors simultaneously at the single-cell level, which remains lacking. Herein, an integrative single-cell secretion analysis platform is presented to profile different secreted factors (four proteins, three extracellular vesicles (EV) phenotypes), spatial distances, and migration information (distances and direction) simultaneously from high-throughput paired single cells using an antibody-barcode microchip. Applying the platform to analyze the tumor-stromal and tumor-immune interactions with the human oral squamous cell carcinoma (OSCC) cell lines and primary OSCC cells reveals that the initial distances between cells would determine their migratory distances and direction to approach stable organization. The cell-cell in close proximity enhances protein secretions while attenuating EV secretions. Migration has a more profound correlation with protein secretions than EV secretions, in which absolute migration distance affects protein secretions significantly but not the direction. These findings highlight the significance of spatial organization in regulating cell signaling behaviors and demonstrate that the integrative single-cell secretion profiling platform is well-suited for a comprehensive dissection of intercellular communication and interactions, providing new avenues for understanding cell-cell interaction biology and how different signaling behaviors coordinate within the tumor microenvironment.