Intestinal-level anti-inflammatory bioactivities of whole wheat: Rationale, design, and methods of a randomized, controlled, crossover dietary trial in adults with prediabetes.

Randomized controlled trials (RCT) demonstrate that whole wheat consumption improves glycemia. However, substantial inter-individual variation is often observed, highlighting that dietary whole grain recommendations may not support the health of all persons. The objective of this report is to describe the rationale and design of a planned RCT aimed at establishing the gut microbiota and metabolome signatures that predict whole wheat-mediated improvements in glucose tolerance in adults with prediabetes. It is hypothesized that a controlled diet containing wheat bread (WHEAT; 160 g/day) compared with refined bread (WHITE) will improve glucose tolerance in a gut microbiota-mediated manner. Biospecimens will be collected before and after each 2-week study arm. Testing for oral glucose tolerance and gastrointestinal permeability will be performed post-intervention. Assessments will include oral glucose tolerance (primary outcome) and secondary outcomes including gut microbiota, targeted and untargeted metabolomics of fecal and plasma samples, intestinal and host inflammatory responses, and intestinal permeability. WHEAT is predicted to alleviate glucose intolerance by shifting microbiota composition to increase short-chain fatty acid-producing bacteria while reducing populations implicated in intestinal inflammation, barrier dysfunction, and systemic endotoxemia. Further, benefits from WHEAT are anticipated to correlate with gut-level and systemic metabolomic responses that can help to explain the expected inter-individual variability in glucose tolerance. Thus, knowledge gained from integrating multi-omic responses associating with glucose tolerance could help to establish a precision nutrition-based framework that can alleviate cardiometabolic risk. This framework could inform novel dietary whole grain recommendations by enhancing our understanding of inter-individual responsiveness to whole grain consumption.

Single-nucleus transcriptional profiling reveals TCF7L2 as a key regulator in adipogenesis in goat skeletal muscle development.

Intramuscular adipogenesis plays an important role in muscle development, which determines the quality of goat meat. However, its underlying cellular and molecular mechanisms remain poorly understood. In this study, we provided detailed cellular atlases of goat longissimus dorsi during muscle development at single-nucleus resolution, and identified the subpopulations of fibroblasts/fibro-adipogenic progenitors (FAPs) and muscle satellite cell (MuSC), as well as the differentiation trajectory of FAPs subpopulations. Cellular ligand-receptor interaction analysis revealed enriched BMP and IGF pathways implicated in within-tissue crosstalk centered around FAPs. Through single-nucleus gene regulatory network analysis and in vitro interference verification, we found that TCF7L2 was a critical transcriptional factor (TF) in early adipogenesis in skeletal muscle. Overall, our work reveals the cellular intricacies and diversity of goat longissimus dorsi during muscle development, implementing insights into the critical roles of BMP, IGF pathways and TCF7L2 TF in intramuscular adipogenesis.

Insecticide-Induced Metabolic Dysregulation in Model Microbe E. coli Discovered by Comprehensive Metabolic Profiling.

Fipronil, malathion, and permethrin are widely used insecticides in agriculture, public areas, and residential spaces. The globally abused application of these chemicals results in residues surpassing established maximum residue levels, giving rise to potential toxicity in unintended organisms. Long-term exposure and the persistent accumulation of these insecticides in animals and humans pose threats such as neurotoxicity, liver and kidney damage, and microbiota dysbiosis. Despite the known risks, the specific impact of these insecticides on gut microbiota and their metabolic processes, as well as the subsequent effects on host health, remain largely unknown. This study aimed to address this gap by utilizing nonpathogenic Escherichia coli as a representative of human gut bacteria and examining its growth and metabolic perturbations induced by exposure to fipronil, malathion, and permethrin. Our research showed that exposure of E. coli to fipronil, malathion, and permethrin at physiologically relevant concentrations resulted in significant growth inhibition. Furthermore, we have observed the biodegradation of fipronil and permethrin by E. coli, while no biodegradation was found for malathion. Thus, E. coli is capable of degrading fipronil and permethrin, thereby enabling the removal of those substances. Next, we studied how insecticides affect bacterial metabolism to understand their influence on the functions of the microbes. Our metabolomics analysis revealed chemical-dependent alterations in metabolic profiles and metabolite compositions following insecticide exposure. These changes encompassed shifts in carboxylic acids and derivatives, organooxygen compounds, as well as indoles and their derivatives. To gain a deeper insight into the systematic changes induced by these insecticides, we conducted a metabolic pathway analysis. Our data indicated that fipronil, compared with malathion and permethrin, exhibited opposite regulation in glycine, serine, and threonine metabolism and valine, leucine, and isoleucine biosynthesis. In summary, our study demonstrates the capability of E. coli to degrade fipronil and permethrin, leading to their removal, while malathion remains unaffected. Additionally, we reveal chemical-dependent alterations in bacterial metabolism induced by insecticide exposure, with specific impacts on metabolic pathways, particularly in pathways related to amino acid metabolism.

New Metabolomic Insights Into Cancer.

ABSTRACT: Cancer has been marked by metabolic irregularities that fuel various aggressive activities such as rapid cell proliferation, evasion of the immune system, and spread to distant organs. Therefore, exploiting cancer metabolism for diagnosis, monitoring, or treatment has been extensively studied in the past couple of decades with various molecular and cellular techniques. More recently, investigating cancer diagnostics and treatments through advanced metabolomics has emerged, and these comprehensive approaches provide a holistic understanding of cancer metabolism, which supported the discovery of metabolic targets relevant across multiple cancer types and the development of more effective treatments. This study offers highlights of new knowledge on cancer metabolism enabled by recent metabolomics studies and their potential applications in aiding cancer research and predicting cancer treatment outcomes. Specifically, we discussed the use of advanced metabolomics in cancer metabolism, tumor microenvironment, and cancer immunotherapy studies to provide valuable insights that can shape future research efforts in the dynamic field of cancer metabolism research.

MiR-196a Promotes Lipid Deposition in Goat Intramuscular Preadipocytes by Targeting MAP3K1 and Activating PI3K-Akt Pathway.

Meat quality in goats is partly determined by the intramuscular fat (IMF) content, which is associated with the proliferation and differentiation of intramuscular preadipocytes. Emerging studies have suggested that miRNA plays a crucial role in adipocyte proliferation and differentiation. In our recent study, we observed the expression variations in miR-196a in the longissimus dorsi muscle of Jianzhou goats at different ages. However, the specific function and underlying mechanism of miR-196a in IMF deposition are still unclear. This study demonstrated that miR-196a significantly enhanced adipogenesis and apoptosis and reduced the proliferation of preadipocytes. Subsequently, RNA-seq was employed to determine genes regulated by miR-196a, and 677 differentially expressed genes were detected after miR-196a overexpression. The PI3K-Akt pathway was identified as activated in miR-196a regulating intramuscular adipogenesis via Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and further verified via Western blot and rescue assays. Lastly, using RT-qPCR, Western blot, dual-luciferase, and rescue assays, we found that miR-196a promoted adipogenesis and suppressed the proliferation of intramuscular preadipocytes by the downregulation of MAP3K1. In summary, these results suggest that miR-196a regulates IMF deposition by targeting MAP3K1 and activating the PI3K-Akt pathway and provide a theoretical foundation for improving goat meat quality through molecular breeding.

An Explainable Graph Neural Framework to Identify Cancer-Associated Intratumoral Microbial Communities.

Microbes are extensively present among various cancer tissues and play critical roles in carcinogenesis and treatment responses. However, the underlying relationships between intratumoral microbes and tumors remain poorly understood. Here, a MIcrobial Cancer-association Analysis using a Heterogeneous graph transformer (MICAH) to identify intratumoral cancer-associated microbial communities is presented. MICAH integrates metabolic and phylogenetic relationships among microbes into a heterogeneous graph representation. It uses a graph transformer to holistically capture relationships between intratumoral microbes and cancer tissues, which improves the explainability of the associations between identified microbial communities and cancers. MICAH is applied to intratumoral bacterial data across 5 cancer types and 5 fungi datasets, and its generalizability and reproducibility are demonstrated. After experimentally testing a representative observation using a mouse model of tumor-microbe-immune interactions, a result consistent with MICAH's identified relationship is observed. Source tracking analysis reveals that the primary known contributor to a cancer-associated microbial community is the organs affected by the type of cancer. Overall, this graph neural network framework refines the number of microbes that can be used for follow-up experimental validation from thousands to tens, thereby helping to accelerate the understanding of the relationship between tumors and intratumoral microbiomes.

Effects of the FHL2 gene on the development of subcutaneous and intramuscular adipocytes in goats.

BACKGROUND: Adipose tissue affects not only the meat quality of domestic animals, but also human health. Adipocyte differentiation is regulated by a series of regulatory genes and cyclins. Four and half-LIM protein (FHL2) is positively correlated with the hypertrophy of adipocytes and can cause symptoms such as obesity and diabetes. RESULT: In the transcriptome sequencing analysis of intramuscular adipocytes after three days of differentiation, the differentially expressed gene FHL2 was found. To further explore the biological significance of the differentially expressed gene FHL2, which was downregulated in the mature adipocytes. We revealed the function of FHL2 in adipogenesis through the acquisition and loss of function of FHL2. The results showed that the overexpression of FHL2 significantly increased the expression of adipogenic genes (PPARγ, C/EBPß) and the differentiation of intramuscular and subcutaneous adipocytes. However, silencing FHL2 significantly inhibited adipocyte differentiation. The overexpression of FHL2 increased the number of adipocytes stained with crystal violet and increased the mRNA expression of proliferation marker genes such as CCNE, PCNA, CCND and CDK2. In addition, it significantly increased the rate of EdU positive cells. In terms of apoptosis, overexpression of FHL2 significantly inhibited the expression of P53 and BAX in both intramuscular and subcutaneous adipocytes, which are involved in cell apoptosis. However, overexpression of FHL2 promoted the expression of BCL, but was rescued by the silencing of FHL2. CONCLUSIONS: In summary, FHL2 may be a positive regulator of intramuscular and subcutaneous adipocyte differentiation and proliferation, and acts as a negative regulator of intramuscular and subcutaneous adipocyte apoptosis. These findings provide a theoretical basis for the subsequent elucidation of FHL2 in adipocytes.

pH-Controlled Chemoselective Rapid Azo-Coupling Reaction (CRACR) Enables Global Profiling of Serotonylation Proteome in Cancer Cells.

Serotonylation has been identified as a novel protein posttranslational modification for decades, where an isopeptide bond is formed between the glutamine residue and serotonin through transamination. Transglutaminase 2 (also known as TGM2 or TGase2) was proven to act as the main "writer" enzyme for this PTM, and a number of key regulatory proteins (including small GTPases, fibronectin, fibrinogen, serotonin transporter, and histone H3) have been characterized as the substrates of serotonylation. However, due to the lack of pan-specific antibodies for serotonylated glutamine, the precise enrichment and proteomic profiling of serotonylation still remain challenging. In our previous research, we developed an aryldiazonium probe to specifically label protein serotonylation in a bioorthogonal manner, which depended on a pH-controlled chemoselective rapid azo-coupling reaction. Here, we report the application of a photoactive aryldiazonium-biotin probe for the global profiling of serotonylation proteome in cancer cells. Thus, over 1,000 serotonylated proteins were identified from HCT 116 cells, many of which are highly related to carcinogenesis. Moreover, a number of modification sites of these serotonylated proteins were determined, attributed to the successful application of our chemical proteomic approach. Overall, these findings provided new insights into the significant association between cellular protein serotonylation and cancer development, further suggesting that to target TGM2-mediated monoaminylation may serve as a promising strategy for cancer therapeutics.

Nuclear Receptor Subfamily 4 Group A Member 1 (NR4A1) Promotes the Adipogenesis of Intramuscular Preadipocytes through PI3K/AKT Pathway in Goats.

As a transcription factor, Nuclear Receptor Subfamily 4 Group A Member 1 (NR4A1) binds to downstream target genes to participate in cell proliferation and cell differentiation. We found that the NR4A1 reached the highest expression at 60 h after the differentiation of goat intramuscular preadipocytes. Overexpression of goat NR4A1 increased the number of intracellular lipid droplets and up-regulated the expression of adipocyte-differentiation-related marker genes including AP2, SREBP1, ACC, GPAM, and DGAT2, while the relative expression levels of Pref-1 and HSL were significantly decreased. On the contrary, after NR4A1 was knocked down by siRNA, the number of intracellular lipid droplets and the relative expression levels of LPL, CEBPα, CEBPß, ACC, and DGAT2 were significantly decreased, and the relative expression levels of Pref-1 and HSL were significantly up-regulated. These results suggest that NR4A1 promotes the differentiation of goat intramuscular preadipocytes. Transcriptome sequencing was carried out after overexpression of goat NR4A1, and the KEGG enrichment analysis result showed that the most differentially expressed genes were related to adipocyte differentiation and were enriched in the PI3K-Akt signaling pathway. LY249002, an inhibitor of the PI3K-Akt signaling pathway, was introduced and decreased the number of intracellular lipid droplets, and the relative expression levels of C/EBPα, SREBP1, AP2, C/EBPß, GPAM, ACC, DGAT1, DGAT2, and ATGL were decreased accordingly. The above results indicate that overexpression of goat NR4A1 may promote the differentiation of intramuscular preadipocytes through the PI3K-Akt signaling pathway.

Subregion-specific transcriptomic profiling of rat brain reveals sex-distinct gene expression impacted by adolescent stress.

Stress during adolescence clearly impacts brain development and function. Sex differences in adolescent stress-induced or exacerbated emotional and metabolic vulnerabilities could be due to sex-distinct gene expression in hypothalamic, limbic, and prefrontal brain regions. However, adolescent stress-induced whole-genome expression changes in key subregions of these brain regions were unclear. In this study, female and male adolescent Sprague Dawley rats received one-hour restraint stress daily from postnatal day (PD) 32 to PD44. Corticosterone levels, body weights, food intake, body composition, and circulating adiposity and sex hormones were measured. On PD44, brain and blood samples were collected. Using RNA-sequencing, sex-specific differences in stress-induced differentially expressed (DE) genes were identified in subregions of the hypothalamus, limbic system, and prefrontal cortex. Canonical pathways reflected well-known sex-distinct maladies and diseases, substantiating the therapeutic potential of the DE genes found in the current study. Thus, we proposed specific sex distinct, adolescent stress-induced transcriptional changes found in the current study as examples of the molecular bases for sex differences witnessed in stress induced or exacerbated emotional and metabolic disorders. Future behavioral studies and single-cell studies are warranted to test the implications of the DE genes identified in this study in sex-distinct stress-induced susceptibilities.

Bariatric Surgery Is Associated with Lower Concentrations of Fecal Secondary Bile Acids and Their Metabolizing Microbial Enzymes: A Pilot Study.

INTRODUCTION: Excess body fat elevates colorectal cancer risk. While bariatric surgery (BRS) induces significant weight loss, its effects on the fecal stream and colon biology are poorly understood. Specifically, limited data exist on the impact of bariatric surgery (BRS) on fecal secondary bile acids (BA), including lithocholic acid (LCA), a putative promotor of colorectal carcinogenesis. METHODS: This cross-sectional case-control study included 44 patients with obesity; 15 pre-BRS (controls) vs. 29 at a median of 24.1 months post-BRS. We examined the fecal concentrations of 11 BA by liquid chromatography and gene abundance of BA-metabolizing bacterial enzymes through fecal metagenomic sequencing. Differences were quantified using non-parametric tests for BA levels and linear discriminant analysis (LDA) effect size (LEfSe) for genes encoding BA-metabolizing enzymes. RESULTS: Total fecal secondary BA concentrations trended towards lower levels post- vs. pre-BRS controls (p = 0.07). Individually, fecal LCA concentrations were significantly lower post- vs. pre-BRS (8477.0 vs. 11,914.0 uM/mg, p < 0.008). Consistent with this finding, fecal bacterial genes encoding BA-metabolizing enzymes, specifically 3-betahydroxycholanate-3-dehydrogenase (EC 1.1.1.391) and 3-alpha-hydroxycholanate dehydrogenase (EC 1.1.1.52), were also lower post- vs. pre-BRS controls (LDA of - 3.32 and - 2.64, respectively, adjusted p < 0.0001). Post-BRS fecal BA concentrations showed significant inverse correlations with weight loss, a healthy diet quality, and increased physical activity. CONCLUSIONS: Concentrations of LCA, a secondary BA, and bacterial genes needed for BA metabolism are lower post-BRS. These changes can impact health and modulate the colorectal cancer cascade. Further research is warranted to examine how surgical alterations and the associated dietary changes impact bile acid metabolism.

Bioorthogonal Labeling and Enrichment of Histone Monoaminylation Reveal Its Accumulation and Regulatory Function in Cancer Cell Chromatin.

Histone monoaminylation (i.e., serotonylation and dopaminylation) is an emerging category of epigenetic mark occurring on the fifth glutamine (Q5) residue of H3 N-terminal tail, which plays significant roles in gene transcription. Current analysis of histone monoaminylation is mainly based on site-specific antibodies and mass spectrometry, which either lacks high resolution or is time-consuming. In this study, we report the development of chemical probes for bioorthogonal labeling and enrichment of histone serotonylation and dopaminylation. These probes were successfully applied for the monoaminylation analysis of in vitro biochemical assays, cells, and tissue samples. The enrichment of monoaminylated histones by the probes further confirmed the crosstalk between H3Q5 monoaminylation and H3K4 methylation. Finally, combining the ex vivo and in vitro analyses based on the developed probes, we have shown that both histone serotonylation and dopaminylation are highly enriched in tumor tissues that overexpress transglutaminase 2 (TGM2) and regulate the three-dimensional architecture of cellular chromatin.

The Tumor Microbiome Reacts to Hypoxia and Can Influence Response to Radiation Treatment in Colorectal Cancer.

Tumor hypoxia has been shown to predict poor patient outcomes in several cancer types, partially because it reduces radiation's ability to kill cells. We hypothesized that some of the clinical effects of hypoxia could also be due to its impact on the tumor microbiome. Therefore, we examined the RNA sequencing data from the Oncology Research Information Exchange Network database of patients with colorectal cancer treated with radiotherapy. We identified microbial RNAs for each tumor and related them to the hypoxic gene expression scores calculated from host mRNA. Our analysis showed that the hypoxia expression score predicted poor patient outcomes and identified tumors enriched with certain microbes such as Fusobacterium nucleatum. The presence of other microbes, such as Fusobacterium canifelinum, predicted poor patient outcomes, suggesting a potential interaction between hypoxia, the microbiome, and radiation response. To experimentally investigate this concept, we implanted CT26 colorectal cancer cells into immune-competent BALB/c and immune-deficient athymic nude mice. After growth, in which tumors passively acquired microbes from the gastrointestinal tract, we harvested tumors, extracted nucleic acids, and sequenced host and microbial RNAs. We stratified tumors based on their hypoxia score and performed a metatranscriptomic analysis of microbial gene expression. In addition to hypoxia-tropic and -phobic microbial populations, analysis of microbial gene expression at the strain level showed expression differences based on the hypoxia score. Thus, hypoxia gene expression scores seem to associate with different microbial populations and elicit an adaptive transcriptional response in intratumoral microbes, potentially influencing clinical outcomes. SIGNIFICANCE: Tumor hypoxia reduces radiotherapy efficacy. In this study, we explored whether some of the clinical effects of hypoxia could be due to interaction with the tumor microbiome. Hypoxic gene expression scores associated with certain microbes and elicited an adaptive transcriptional response in others that could contribute to poor clinical outcomes.

Chemical Proteomic Profiling of Protein Dopaminylation in Colorectal Cancer Cells.

Histone dopaminylation is a newly identified epigenetic mark that plays a role in the regulation of gene transcription, where an isopeptide bond is formed between the fifth amino acid of H3 (i.e., glutamine) and dopamine. Recently, we developed a chemical probe to specifically label and enrich histone dopaminylation via bioorthogonal chemistry. Given this powerful tool, we found that histone H3 glutamine 5 dopaminylation (H3Q5dop) was highly enriched in colorectal tumors, which could be attributed to the high expression level of its regulator, transglutaminase 2 (TGM2), in colon cancer cells. Due to the enzyme promiscuity of TGM2, nonhistone proteins have also been identified as dopaminylation targets; however, the dopaminylated proteome in cancer cells still remains elusive. Here, we utilized our chemical probe to enrich dopaminylated proteins from colorectal cancer cells in a bioorthogonal manner and performed the chemical proteomics analysis. Therefore, 425 dopaminylated proteins were identified, many of which are involved in nucleic acid metabolism and transcription pathways. More importantly, a number of dopaminylation sites were identified and attributed to the successful application of our chemical probe. Overall, these findings shed light on the significant association between cellular protein dopaminylation and cancer development, further suggesting that targeting these pathways may become a promising anticancer strategy.

LC-HRMS-based metabolomics and lipidomics analyses of a novel probiotic Akkermansia Muciniphila in response to different nutritional stimulations.

The mucin-degrading gut commensal Akkermansia muciniphila (A. muciniphila) negatively correlates with various diseases, including metabolic disorders, neurodegenerative disorders, and cancers, through interacting with host receptors by diverse molecules. Still, their exact metabolic capability within the nutrient-rich environment (such as in the human gut) is not fully characterized. Therefore, in the present study, we investigated the comprehensive metabolome and lipidome of A. muciniphila after supplementation of four major gut microbial nutrients: mucin, inorganic salts, bile salts, and short-chain fatty acids (SCFAs). Our results showed that mucin is the predominant driver of the different lipidomic and metabolomic profiles of A. muciniphila, and it promotes the overall growth of this bacteria. While the addition of inorganic salts, bile salts, and SCFAs was found to inhibit the growth of A. muciniphila. Interestingly, inorganic salts affected the purine metabolism in A. muciniphila cultures, while adding bile salts significantly increased the production of other bile acids and N-acyl amides. Lastly, SCFAs were identified to alter the A. muciniphila energy utilization of triglycerides, fatty acyls, and phosphatidylethanolamines. To our knowledge, this is the first study to examine the comprehensive lipidome and metabolome of A. muciniphila, which highlights the importance of nutritional impacts on the lipidome and metabolome of A. muciniphila and hence providing foundational knowledge to unveil the potential effects of A. muciniphila on host health.

Arachidonic Acid Mobilization and Peroxidation Promote Microglial Dysfunction in Aß Pathology.

Aberrant increase of arachidonic acid (ARA) has long been implicated in the pathology of Alzheimer's disease (AD), while the underlying causal mechanism remains unclear. In this study, we revealed a link between ARA mobilization and microglial dysfunction in Aß pathology. Lipidomic analysis of primary microglia from AppNL-GF mice showed a marked increase in free ARA and lysophospholipids (LPLs) along with a decrease in ARA-containing phospholipids, suggesting increased ARA release from phospholipids (PLs). To manipulate ARA-containing PLs in microglia, we genetically deleted lysophosphatidylcholine acyltransferase 3 (Lpcat3), the main enzyme catalyzing the incorporation of ARA into PLs. Loss of microglial Lpcat3 reduced the levels of ARA-containing PLs, free ARA and LPLs, leading to a compensatory increase in monounsaturated fatty acid (MUFA)-containing PLs in both male and female App NL-GF mice. Notably, the reduction of ARA in microglia significantly ameliorated oxidative stress and inflammatory responses while enhancing the phagocytosis of Aß plaques and promoting the compaction of Aß deposits. Mechanistically, scRNA seq suggested that LPCAT3 deficiency facilitates phagocytosis by facilitating de novo lipid synthesis while protecting microglia from oxidative damage. Collectively, our study reveals a novel mechanistic link between ARA mobilization and microglial dysfunction in AD. Lowering brain ARA levels through pharmacological or dietary interventions may be a potential therapeutic strategy to slow down AD progression.

Chemical proteomic profiling of protein dopaminylation in colorectal cancer cells.

Histone dopaminylation is a newly identified epigenetic mark that plays a role in the regulation of gene transcription, where an isopeptide bond is formed between the fifth amino acid residue of H3 ( i.e. , glutamine) and dopamine. In our previous studies, we discovered that the dynamics of this post-translational modification (including installation, removal, and replacement) were regulated by a single enzyme, transglutaminase 2 (TGM2), through reversible transamination. Recently, we developed a chemical probe to specifically label and enrich histone dopaminylation via bioorthogonal chemistry. Given this powerful tool, we found that histone H3 glutamine 5 dopaminylation (H3Q5dop) was highly enriched in colorectal tumors, which could be attributed to the high expression level of TGM2 in colon cancer cells. Due to the enzyme promiscuity of TGM2, non-histone proteins have also been identified as targets of dopaminylation on glutamine residues, however, the dopaminylated proteome in cancer cells still remains elusive. Here, we utilized our chemical probe to enrich dopaminylated proteins from colorectal cancer cells in a bioorthogonal manner and performed the chemical proteomics analysis. Therefore, 425 dopaminylated proteins were identified, many of which are involved in nucleic acid metabolism and transcription pathways. More importantly, a number of modification sites of these dopaminylated proteins were identified, attributed to the successful application of our chemical probe. Overall, these findings shed light on the significant association between cellular protein dopaminylation and cancer development, further suggesting that to block the installation of protein dopaminylation may become a promising anti-cancer strategy.

pH-Controlled chemoselective rapid azo-coupling reaction (CRACR) enables global profiling of serotonylation proteome in cancer cells.

Serotonylation has been identified as a novel protein post-translational modification for decades, where an isopeptide bond is formed between the glutamine residue and serotonin through transamination. Transglutaminase 2 (also known as TGM2 or TGase2) was proven to act as the main writer enzyme for this PTM and a number of key regulatory proteins (including small GTPases, fibronectin, fibrinogen, serotonin transporter, and histone H3) have been characterized as the substrates of serotonylation. However, due to the lack of pan-specific antibodies for serotonylated glutamine, the precise enrichment and proteomic profiling of serotonylation still remain challenging. In our previous research, we developed an aryldiazonium probe to specifically label protein serotonylation in a bioorthogonal manner, which depended on a pH-controlled chemoselective rapid azo-coupling reaction (CRACR). Here, we report the application of a photoactive aryldiazonium-biotin probe for the global profiling of serotonylation proteome in cancer cells. Thus, over 1,000 serotonylated proteins were identified from HCT 116 cells, many of which are highly related to carcinogenesis. Moreover, a number of modification sites of these serotonylated proteins were determined, attributed to the successful application of our chemical proteomic approach. Overall, these findings provided new insights into the significant association between cellular protein serotonylation and cancer development, further suggesting that to target TGM2-mediated monoaminylation may serve as a promising strategy for cancer therapeutics.

Bioorthogonal labeling and enrichment of histone monoaminylation reveal its accumulation and regulatory function in cancer cell chromatin.

Histone monoaminylation ( i . e ., serotonylation and dopaminylation) is an emerging category of epigenetic mark occurring on the fifth glutamine (Q5) residue of H3 N-terminal tail, which plays significant roles in gene transcription. Current analysis of histone monoaminylation is mainly based on site-specific antibodies and mass spectrometry, which either lacks high resolution or is time-consuming. In this study, we report the development of chemical probes for bioorthogonal labeling and enrichment of histone serotonylation and dopaminylation. These probes were successfully applied for the monoaminylation analysis of in vitro biochemical assays, cells, and tissue samples. The enrichment of monoaminylated histones by the probes further confirmed the crosstalk between H3Q5 monoaminylation and H3K4 methylation. Finally, combining the ex vivo and in vitro analyses based on the developed probes, we have shown that both histone serotonylation and dopaminylation are highly enriched in tumor tissues that overexpress transglutaminase 2 (TGM2) and regulate the three-dimensional architecture of cellular chromatin.

Crosstalk between innate immunity and rumen-fecal microbiota under the cold stress in goats.

The balance of the microbiome, which is sensitive to temperature changes, plays a crucial role in maintaining overall health and reducing the risk of diseases. However, the specific mechanisms by which immunity and microbiota interact to adapt to cold stress have yet to be addressed. In this study, Nanjiang Yellow goats were chosen as a model and sampled during the cold (winter, cold stress) and warm (spring) seasons, respectively. Analyses of serum immune factors, as well as the composition of rumen and fecal microbial communities, were conducted to explore the crosstalk between microbiota and innate immunity under cold stress. Significantly increased levels of IgA (P < 0.01) were observed in the cold season compared to the warm season. Conversely, the levels of IL-2 (P = 0.02) and IL-6 (P < 0.01) diminished under cold stress. However, no significant differences were observed in IgG (P = 0.89), IgM (P = 0.42), and IL-4 (P = 0.56). While there were no significant changes in the diversity of bacterial communities between the warm and cold seasons, positive correlations between serum IgA, IL-2, IL-6 concentrations and several genera were observed. Furthermore, the weighted gene co-expression network analysis indicated that the microbiota enriched in the MEbrown module positively correlated with IgA, while the microbiota enriched in the MEblue module positively correlated with IL-2 and IL-6. The strong correlation between certain probiotics, including Alistipes, Bacteroides, Blautia, and Prevotellaceae_UCG.004, and the concentration of IL-2, and IL-6 suggests their potential role in immunomodulatory properties. This study provides valuable insights into the crosstalk between microbial communities and immune responses under the challenge of cold stress. Further studies on the immunomodulatory properties of these probiotics would contribute to the development of strategies to enhance the stress resistance of animals for improved overall health and survival.