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Cell Discovery Jun 2023Alpha-fetoprotein (AFP)-secreting hepatocellular carcinoma (HCC), which accounts for ~75% of HCCs, is more aggressive with a worse prognosis than those without AFP...
Alpha-fetoprotein (AFP)-secreting hepatocellular carcinoma (HCC), which accounts for ~75% of HCCs, is more aggressive with a worse prognosis than those without AFP production. The mechanism through which the interaction between tumors and the microenvironment leads to distinct phenotypes is not yet clear. Therefore, our study aims to identify the characteristic features and potential treatment targets of AFP-negative HCC (ANHC) and AFP-positive HCC (APHC). We utilized single-cell RNA sequencing to analyze 6 ANHC, 6 APHC, and 4 adjacent normal tissues. Integrated multi-omics analysis together with survival analysis were also performed. Further validation was conducted via cytometry time-of-flight on 30 HCCs and multiplex immunohistochemistry on additional 59 HCCs. Our data showed that the genes related to antigen processing and interferon-γ response were abundant in tumor cells of APHC. Meanwhile, APHC was associated with multifaceted immune distortion, including exhaustion of diverse T cell subpopulations, and the accumulation of tumor-associated macrophages (TAMs). Notably, TAM-SPP1 was highly enriched in APHC, as was its receptor CD44 on T cells and tumor cells. Targeting the Spp1-Cd44 axis restored T cell function in vitro and significantly reduced tumor burden when treated with either anti-Spp1 or anti-Cd44 antibody alone or in combination with anti-Pd-1 antibody in the mouse model. Furthermore, elevated IL6 and TGF-β1 signaling contributed to the enrichment of TAM-SPP1 in APHC. In conclusion, this study uncovered a highly suppressive microenvironment in APHC and highlighted the role of TAM-SPP1 in regulating the immune microenvironment, thereby revealing the SPP1-CD44 axis as a promising target for achieving a more favorable immune response in APHC treatment.
PubMed: 37336873
DOI: 10.1038/s41421-023-00563-x -
Journal of Molecular Cell Biology Nov 2023Chemoresistance is a primary cause of treatment failure in pancreatic cancer. Identifying cell surface markers specifically expressed in chemoresistant cancer cells...
Chemoresistance is a primary cause of treatment failure in pancreatic cancer. Identifying cell surface markers specifically expressed in chemoresistant cancer cells (CCCs) could facilitate targeted therapies to overcome chemoresistance. We performed an antibody-based screen and found that TRA-1-60 and TRA-1-81, two 'stemness' cell surface markers, are highly enriched in CCCs. Furthermore, TRA-1-60+/TRA-1-81+ cells are chemoresistant compared to TRA-1-60-/TRA-1-81- cells. Transcriptome profiling identified UGT1A10, shown to be both necessary and sufficient to maintain TRA-1-60/TRA-1-81 expression and chemoresistance. From a high-content chemical screen, we identified Cymarin, which downregulates UGT1A10, eliminates TRA-1-60/TRA-1-81 expression, and increases chemosensitivity both in vitro and in vivo. Finally, TRA-1-60/TRA-1-81 expression is highly specific in primary cancer tissue and positively correlated with chemoresistance and short survival, which highlights their potentiality for targeted therapy. Therefore, we discovered a novel CCC surface marker regulated by a pathway that promotes chemoresistance, as well as a leading drug candidate to target this pathway.
Topics: Humans; Cell Line, Tumor; Drug Resistance, Neoplasm; Pancreatic Neoplasms; Gene Expression Profiling
PubMed: 37327088
DOI: 10.1093/jmcb/mjad039 -
Nature Jun 2023The immune phenotype of a tumour is a key predictor of its response to immunotherapy. Patients who respond to checkpoint blockade generally present with immune-inflamed...
The immune phenotype of a tumour is a key predictor of its response to immunotherapy. Patients who respond to checkpoint blockade generally present with immune-inflamed tumours that are highly infiltrated by T cells. However, not all inflamed tumours respond to therapy, and even lower response rates occur among tumours that lack T cells (immune desert) or that spatially exclude T cells to the periphery of the tumour lesion (immune excluded). Despite the importance of these tumour immune phenotypes in patients, little is known about their development, heterogeneity or dynamics owing to the technical difficulty of tracking these features in situ. Here we introduce skin tumour array by microporation (STAMP)-a preclinical approach that combines high-throughput time-lapse imaging with next-generation sequencing of tumour arrays. Using STAMP, we followed the development of thousands of arrayed tumours in vivo to show that tumour immune phenotypes and outcomes vary between adjacent tumours and are controlled by local factors within the tumour microenvironment. Particularly, the recruitment of T cells by fibroblasts and monocytes into the tumour core was supportive of T cell cytotoxic activity and tumour rejection. Tumour immune phenotypes were dynamic over time and an early conversion to an immune-inflamed phenotype was predictive of spontaneous or therapy-induced tumour rejection. Thus, STAMP captures the dynamic relationships of the spatial, cellular and molecular components of tumour rejection and has the potential to translate therapeutic concepts into successful clinical strategies.
Topics: Humans; Immunotherapy; Neoplasms; T-Lymphocytes; Tumor Microenvironment; Phenotype; Fibroblasts; Monocytes; Immune Checkpoint Inhibitors
PubMed: 37258670
DOI: 10.1038/s41586-023-06132-2 -
Scientific Reports Sep 2023FUS-ERG is a chimeric gene with a poor prognosis, found in myelodysplastic syndromes (MDS) and acute myeloid leukaemia (AML). It remains unclear whether DNA...
FUS-ERG is a chimeric gene with a poor prognosis, found in myelodysplastic syndromes (MDS) and acute myeloid leukaemia (AML). It remains unclear whether DNA hypomethylating agents, including azacitidine (Aza), are effective in FUS-ERG-harbouring AML and how FUS-ERG induces chemoresistance. Stable Ba/F3 transfectants with FUS-ERG were repeatedly exposed to Aza for 7 days of treatment and at 21-day intervals to investigate Aza sensitivity. Stable FUS-ERG transfectants acquired resistance acquired resistance after three courses of Aza exposure. RNA sequencing (RNA-seq) was performed when Aza susceptibility began to change; genes with altered expression or transcript variants were identified. Molecular signatures of these genes were analysed using gene ontology. RNA-seq analyses identified 74 upregulated and 320 downregulated genes involved in cell motility, cytokine production, and kinase activity. Additionally, 1321 genes with altered transcript variants were identified, revealing their involvement in chromatin organisation. In a clinical case of AML with FUS-ERG, we compared whole-genome alterations between the initial MDS diagnosis and AML recurrence after Aza treatment. Genes with non-synonymous or near mutations in transcription regulatory areas (TRAs), additionally detected in AML recurrence, were collated with the gene list from RNA-seq to identify genes involved in acquiring Aza resistance in the presence of FUS-ERG. Whole-genome sequencing of clinical specimens identified 29 genes with non-synonymous mutations, including BCOR, and 48 genes located within 20 kb of 54 TRA mutations in AML recurrence. These genes were involved in chromatin organisation and included NCOR2 as an overlapping gene with RNA-seq data. Transcription regulators involved in mutated TRAs were skewed and included RCOR1 in AML recurrence. We tested the efficacy of BH3 mimetics, including venetoclax and S63845, in primary Aza-resistant AML cells treated with FUS-ERG. Primary FUS-ERG-harbouring AML cells acquiring Aza resistance affected the myeloid cell leukaemia-1 (MCL1) inhibitor S63845 but not while using venetoclax, despite no mutations in BCL2. FUS-ERG promoted Aza resistance after several treatments. The disturbance of chromatin organisation might induce this by co-repressors, including BCOR, NCOR2, and RCOR1. MCL1 inhibition could partially overcome Aza resistance in FUS-ERG-harbouring AML cells.
Topics: Azacitidine; Myeloid Cell Leukemia Sequence 1 Protein; Chromatin
PubMed: 37660196
DOI: 10.1038/s41598-023-41230-1 -
International Journal of Molecular... Feb 2024Rosavin, a phenylpropanoid in 's rhizome, and an adaptogen, is known for enhancing the body's response to environmental stress. It significantly affects cellular... (Review)
Review
Rosavin, a phenylpropanoid in 's rhizome, and an adaptogen, is known for enhancing the body's response to environmental stress. It significantly affects cellular metabolism in health and many diseases, particularly influencing bone tissue metabolism. In vitro, rosavin inhibits osteoclastogenesis, disrupts F-actin ring formation, and reduces the expression of osteoclastogenesis-related genes such as cathepsin K, calcitonin receptor (CTR), tumor necrosis factor receptor-associated factor 6 (TRAF6), tartrate-resistant acid phosphatase (TRAP), and matrix metallopeptidase 9 (MMP-9). It also impedes the nuclear factor of activated T-cell cytoplasmic 1 (NFATc1), c-Fos, the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and mitogen-activated protein kinase (MAPK) signaling pathways and blocks phosphorylation processes crucial for bone resorption. Moreover, rosavin promotes osteogenesis and osteoblast differentiation and increases mouse runt-related transcription factor 2 (Runx2) and osteocalcin (OCN) expression. In vivo studies show its effectiveness in enhancing bone mineral density (BMD) in postmenopausal osteoporosis (PMOP) mice, restraining osteoclast maturation, and increasing the active osteoblast percentage in bone tissue. It modulates mRNA expressions by increasing eukaryotic translation elongation factor 2 (EEF2) and decreasing histone deacetylase 1 (HDAC1), thereby activating osteoprotective epigenetic mechanisms, and alters many serum markers, including decreasing cross-linked C-telopeptide of type I collagen (CTX-1), tartrate-resistant acid phosphatase 5b (TRACP5b), receptor activator for nuclear factor κ B ligand (RANKL), macrophage-colony-stimulating factor (M-CSF), and TRAP, while increasing alkaline phosphatase (ALP) and OCN. Additionally, when combined with zinc and probiotics, it reduces pro-osteoporotic matrix metallopeptidase 3 (MMP-3), interleukin 6 (IL-6), and tumor necrosis factor α (TNF-α), and enhances anti-osteoporotic interleukin 10 (IL-10) and tissue inhibitor of metalloproteinase 3 (TIMP3) expressions. This paper aims to systematically review rosavin's impact on bone tissue metabolism, exploring its potential in osteoporosis prevention and treatment, and suggesting future research directions.
Topics: Animals; Mice; Osteoclasts; Tartrate-Resistant Acid Phosphatase; Osteogenesis; Bone Resorption; Cell Differentiation; NF-kappa B; Metalloproteases; RANK Ligand; NFATC Transcription Factors; Disaccharides
PubMed: 38396794
DOI: 10.3390/ijms25042117 -
Journal, Genetic Engineering &... Nov 2023Early childhood caries is a significant public health concern affecting about 600 million children globally. The etiology of early childhood caries can be explained as... (Review)
Review
BACKGROUND
Early childhood caries is a significant public health concern affecting about 600 million children globally. The etiology of early childhood caries can be explained as an interplay between genetic and environmental factors. Single nucleotide polymorphisms are the most common variations in the human genome. Genetic variations of immune response genes can modify the defense response of the host, and alter the susceptibility to bacterial colonization of the oral cavity and early childhood caries. The aim of this systematic review is to identify genetic variants of immune response genes associated with early childhood caries.
RESULTS
A total of 7124 articles were identified by conducting an elaborate search across various electronic databases and genome-wide association studies databases. Subsequent to exclusion at various stages, fifteen articles qualified to be included into the present review. Risk of bias assessment was done with the Q-genie tool. Quantitative synthesis revealed that the odds ratio for TT and CC genotypes of rs11362 was 1.07 (0.67-1.71) and 1.16 (0.84-1.60), respectively. Gene-based analysis revealed a statistically significant association between variants of tumor necrosis factor-alpha gene and T-cell receptor alpha variable 4 locus with early childhood caries. Gene clustering showed the presence of three functional clusters. To comprehend the protein-protein interaction, the bioinformatic tool of "Search Tools for the Retrieval of Interacting Genes and Proteins" was used. Among the biological processes and the reactome pathways, complement activation through the lectin pathway showed the highest strength of association with early childhood caries. To understand the interaction and functionality of the genes, "gene function prediction using Multiple Association Network Integration Algorithm" was used, which revealed that the genes were linked by physical interaction (39.34%) and through co-expression (34.88%).
CONCLUSIONS
Genotype TT of rs7217186 of arachidonate 15-lipoxygenase gene was a risk factor for early childhood caries. Multiple genetic variants of T-cell receptor alpha variable 4 locus and tumor necrosis factor-alpha gene were associated with increased susceptibility to early childhood caries. Polymorphisms of genes regulating the lectin pathway of complement activation can modify the susceptibility to early childhood caries.
PubMed: 37971556
DOI: 10.1186/s43141-023-00566-x -
BioRxiv : the Preprint Server For... Nov 2023Allelic variability in the adaptive immune receptor loci, which harbor the gene segments that encode B cell and T cell receptors (BCR/TCR), has been shown to be of...
Allelic variability in the adaptive immune receptor loci, which harbor the gene segments that encode B cell and T cell receptors (BCR/TCR), has been shown to be of critical importance for immune responses to pathogens and vaccines. In recent years, B cell and T cell receptor repertoire sequencing (Rep-Seq) has become widespread in immunology research making it the most readily available source of information about allelic diversity in immunoglobulin (IG) and T cell receptor (TR) loci in different populations. Here we present a novel algorithm for extra-sensitive and specific variable (V) and joining (J) gene allele inference and genotyping allowing reconstruction of individual high-quality gene segment libraries. The approach can be applied for inferring allelic variants from peripheral blood lymphocyte BCR and TCR repertoire sequencing data, including hypermutated isotype-switched BCR sequences, thus allowing high-throughput genotyping and novel allele discovery from a wide variety of existing datasets. The developed algorithm is a part of the MiXCR software ( https://mixcr.com ) and can be incorporated into any pipeline utilizing upstream processing with MiXCR. We demonstrate the accuracy of this approach using Rep-Seq paired with long-read genomic sequencing data, comparing it to a widely used algorithm, TIgGER. We applied the algorithm to a large set of IG heavy chain (IGH) Rep-Seq data from 450 donors of ancestrally diverse population groups, and to the largest reported full-length TCR alpha and beta chain (TRA; TRB) Rep-Seq dataset, representing 134 individuals. This allowed us to assess the genetic diversity of genes within the IGH, TRA and TRB loci in different populations and demonstrate the connection between antibody repertoire gene usage and the number of allelic variants present in the population. Finally we established a database of allelic variants of V and J genes inferred from Rep-Seq data and their population frequencies with free public access at https://vdj.online .
PubMed: 38014266
DOI: 10.1101/2023.10.10.561703 -
Insects Jul 2023In the model system for genetics, , sexual differentiation and male courtship behavior are controlled by sex-specific splicing of () and (). In vitro and in vivo...
In the model system for genetics, , sexual differentiation and male courtship behavior are controlled by sex-specific splicing of () and (). In vitro and in vivo studies showed that female-specific Transformer (TRA) and the non-sex-specific Transformer 2 (TRA2) splicing factors interact, forming a complex promoting and female-specific splicing. TRA/TRA2 complex binds to 13 nt long sequence repeats in their pre-mRNAs. In the Mediterranean fruitfly (Medfly), a major agricultural pest, which shares with Drosophila a ~120 million years old ancestor, and genes seem to promote female-specific splicing of and , which contain conserved TRA/TRA2 binding repeats. Unlike Drosophila , autoregulates its female-specific splicing through these putative regulatory repeats. Here, a yeast two-hybrid assay shows that CcTRA interacts with CcTRA2, despite its high amino acid divergence compared to Drosophila TRA. Interestingly, CcTRA2 interacts with itself, as also observed for Drosophila TRA2. We also generated a three-dimensional model of the complex formed by CcTRA and CcTRA2 using predictive approaches based on Artificial Intelligence. This structure also identified an evolutionary and highly conserved putative TRA2 recognition motif in the TRA sequence. The Y2H approach, combined with powerful predictive tools of three-dimensional protein structures, could use helpful also in this and other insect species to understand the potential links between different upstream proteins acting as primary sex-determining signals and the conserved TRA and TRA2 transducers.
PubMed: 37504611
DOI: 10.3390/insects14070605 -
Microbiology Spectrum Jun 2023Bacterial conjugation is one of the most abundant horizontal gene transfer (HGT) mechanisms, playing a fundamental role in prokaryote evolution. A better understanding...
Bacterial conjugation is one of the most abundant horizontal gene transfer (HGT) mechanisms, playing a fundamental role in prokaryote evolution. A better understanding of bacterial conjugation and its cross talk with the environment is needed for a more complete understanding of HGT mechanisms and to fight the dissemination of malicious genes between bacteria. Here, we studied the effect of outer space, microgravity, and additional key environmental cues on transfer ( gene expression and conjugation efficiency, using the under studied broad-host range plasmid pN3, as a model. High resolution scanning electron microscopy revealed the morphology of the pN3 conjugative pili and mating pair formation during conjugation. Using a nanosatellite carrying a miniaturized lab, we studied pN3 conjugation in outer space, and used qRT-PCR, Western blotting and mating assays to determine the effect of ground physicochemical parameters on gene expression and conjugation. We showed for the first time that bacterial conjugation can occur in outer space and on the ground, under microgravity-simulated conditions. Furthermore, we demonstrated that microgravity, liquid media, elevated temperature, nutrient depletion, high osmolarity and low oxygen significantly reduce pN3 conjugation. Interestingly, under some of these conditions we observed an inverse correlation between gene transcription and conjugation frequency and found that induction of at least and can negatively affect pN3 conjugation frequency in a dose-dependent manner. Collectively, these results uncover pN3 regulation by various environmental cues and highlight the diversity of conjugation systems and the different ways in which they may be regulated in response to abiotic signals. Bacterial conjugation is a highly ubiquitous and promiscuous process, by which a donor bacterium transfers a large portion of genetic material to a recipient cell. This mechanism of horizontal gene transfer plays an important role in bacterial evolution and in the ability of bacteria to acquire resistance to antimicrobial drugs and disinfectants. Bacterial conjugation is a complex and energy-consuming process, that is tightly regulated and largely affected by various environmental signals sensed by the bacterial cell. Comprehensive knowledge about bacterial conjugation and the ways it is affected by environmental cues is required to better understand bacterial ecology and evolution and to find new effective ways to counteract the threating dissemination of antibiotic resistance genes between bacterial populations. Moreover, characterizing this process under stress or suboptimal growth conditions such as elevated temperatures, high salinity or in the outer space, may provide insights relevant to future habitat environmental conditions.
Topics: Conjugation, Genetic; Cues; Plasmids; Bacteria; Gene Transfer, Horizontal
PubMed: 36995224
DOI: 10.1128/spectrum.03688-22 -
Nucleic Acids Research Sep 2023The generation of highly diverse antigen receptors in T and B lymphocytes relies on V(D)J recombination. The enhancer Eα has been implicated in regulating the...
The generation of highly diverse antigen receptors in T and B lymphocytes relies on V(D)J recombination. The enhancer Eα has been implicated in regulating the accessibility of Vα and Jα genes through long-range interactions during rearrangements of the T-cell antigen receptor gene Tcra. However, direct evidence for Eα physically mediating the interaction of Vα and Jα genes is still lacking. In this study, we utilized the 3C-HTGTS assay, a chromatin interaction technique based on 3C, to analyze the higher order chromatin structure of the Tcra locus. Our analysis revealed the presence of sufficient information in the 3C-HTGTS data to detect multiway contacts. Three-way contact analysis of the Tcra locus demonstrated the co-occurrence of the proximal Jα genes, Vα genes and Eα in CD4+CD8+ double-positive thymocytes. Notably, the INT2-TEAp loop emerged as a prominent structure likely to be responsible for bringing the proximal Jα genes and the Vα genes into proximity. Moreover, the enhancer Eα utilizes this loop to establish physical proximity with the proximal Vα gene region. This study provides insights into the higher order chromatin structure of the Tcra locus, shedding light on the spatial organization of chromatin and its impact on V(D)J recombination.
Topics: Chromatin; Receptors, Antigen, T-Cell; Receptors, Antigen, T-Cell, alpha-beta; Thymocytes; V(D)J Recombination; Animals; Mice
PubMed: 37534534
DOI: 10.1093/nar/gkad641