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PubMed2005

Inferring gene transcriptional modulatory relations: a genetical genomics approach.

Bayesian network modeling is a promising approach to define and evaluate gene expression circuits in diverse tissues and cell types under different experimental conditions. The power and practicality of this approach can be improved by restricting the number of potential interactions among genes and by defining causal relations before evaluating posterior probabilities for billions of networks. A newly developed genetical genomics method that combines transcriptome profiling with complex trait analysis now provides strong constraints on network architecture. This method detects those chromosomal intervals responsible for differences in mRNA expression using quantitative trait locus (QTL) mapping. We have developed an efficient Bayesian approach that exploits the genetical genomics method to focus computational effort on the most plausible gene modulatory networks. We exploit a dense marker map for a genetic reference population (GRP) that consists of 32 BXD strains of mice made by intercrossing two progenitor strains--C57BL/6J and DBA/2J. These progenitors differ at approximately 1.3 million known single nucleotide polymorphisms (SNPs), all of which can be exploited to estimate the probability that a gene contains functional polymorphisms that segregate within the GRP. We constructed 66 candidate networks that include all the candidate modulator genes located in the 209 statistically significant trans-acting QTL regions. SNPs that distinguish between the two progenitor strains were used to further winnow the list of candidate modulators. Bayesian network was then used to identify the genetic modulatory relations that best explain the microarray data.

Algorithms↗

PubMed2005

Transcriptome and selected metabolite analyses reveal multiple points of ethylene control during tomato fruit development.

Transcriptome profiling via cDNA microarray analysis identified 869 genes that are differentially expressed in developing tomato (Solanum lycopersicum) pericarp. Parallel phenotypic and targeted metabolite comparisons were employed to inform the expression analysis. Transcript accumulation in tomato fruit was observed to be extensively coordinated and often completely dependent on ethylene. Mutation of an ethylene receptor (Never-ripe [Nr]), which reduces ethylene sensitivity and inhibits ripening, alters the expression of 37% of these 869 genes. Nr also influences fruit morphology, seed number, ascorbate accumulation, carotenoid biosynthesis, ethylene evolution, and the expression of many genes during fruit maturation, indicating that ethylene governs multiple aspects of development both prior to and during fruit ripening in tomato. Of the 869 genes identified, 628 share homology (E-value < or = 1 x 10(-10)) with known gene products or known protein domains. Of these 628 loci, 72 share homology with previously described signal transduction or transcription factors, suggesting complex regulatory control. These results demonstrate multiple points of ethylene regulatory control during tomato fruit development and provide new insights into the molecular basis of ethylene-mediated ripening.

Ascorbic Acid↗

PubMed2026

Integrative Analysis Uncover the Effects and Multi-Omics Features of Thigh Muscle Fat Infiltration.

The health impacts and underlying biological pathways of thigh muscle fat infiltration (TMFI) remain incompletely understood. In this study, we analyzed TMFI measured by magnetic resonance imaging in 55,120 UK Biobank participants and found that higher TMFI was significantly associated with all-cause mortality as well as with all major system-specific diseases examined (p values ranged from 2.50 × 10-88 to 9.97 × 10-04). TMFI also mediated the effects of lifestyle factors on health-related outcomes, with mediation proportions ranging from 6.7% to 71.7%. A genome-wide association study (GWAS) identified 79 lead single nucleotide polymorphisms (SNPs) linked to TMFI, and the polygenic risk score for TMFI was significantly associated with mortality and all incident diseases across examined organ systems in an independent subset of UK Biobank participants of European ancestry who were not included in the TMFI GWAS (n = 362,286, all p < 0.05). Gene-drug interactions identified multiple drugs that could potentially modulate TMFI. Analysis of single-cell transcriptomic data indicated that myogenic cells were strongly linked to TMFI (p = 7.08 × 10-08). Summary-data-based Mendelian randomization and Transcriptome-Wide Association Study analyses revealed numerous genes whose expression in specific tissues was associated with TMFI. Proteomic and metabolomic profiling uncovered a broad array of circulating biomarkers associated with TMFI, many of which mediated the effects of modifiable factors and genetic risk on TMFI. Overall, our results highlight the biological relevance of TMFI to human health and provide insights into the multi-omics mechanisms underlying TMFI, identifying potential targets for interventions.

Humans↗

PubMed2025

The chloroplast 16S rRNA dimethyltransferase BrPFC1 is required for Brassica rapa development under chilling stress.

Chloroplast ribosomal RNA (Ch-rRNA) methylation is critical for plant development and response to low temperatures. Several Ch-rRNA methyltransferases and their catalytic modes, as well as biological relevance, have been reported in model plant species. However, Ch-rRNA methyltransferases and their functional significance remain poorly characterized in crops, including leafy vegetables such as Chinese cabbage. In this study, we screened an EMS-mutagenized Chinese cabbage population and identified a yellow inner leaf (yif) mutant. This mutant develops yellowing inner leaves with reduced chlorophyll accumulation and ultrastructure-impaired chloroplasts under low-temperature conditions. Genetic analysis revealed a premature termination mutation in BrPFC1, encoding the chloroplast-localized 16S rRNA dimethyltransferase. The BrPFC1 mutation (yif) disrupts the dimethylation of 16S rRNA. The cold-sensitive phenotype of the yif mutant can be explained by temperature-dependent defects in the maturation and assembly of chloroplast ribosomes at 4°C. Through integrated analysis of chloroplast and nuclear transcriptomes coupled with translational profiling at 25°C and 4°C, we established that low temperature preferentially upregulates transcripts encoding nuclear-derived ribosomal proteins, while defective 16S rRNA specifically compromises the translational efficiency of chloroplast-encoded photosynthetic complex and ribosomal protein at 4°C. These findings establish rRNA modification by BrPFC1 as a critical regulatory layer for optimizing chloroplast translational efficiency at 4°C, providing mechanistic insights into post-translational adaptation strategies in Chinese cabbage.

Chloroplasts↗

PubMed2005

Characterization of a Corynebacterium glutamicum lactate utilization operon induced during temperature-triggered glutamate production.

Gene expression changes of glutamate-producing Corynebacterium glutamicum were identified in transcriptome comparisons by DNA microarray analysis. During glutamate production induced by a temperature shift, C. glutamicum strain 2262 showed significantly higher mRNA levels of the NCgl2816 and NCgl2817 genes than its non-glutamate-producing derivative 2262NP. Reverse transcription-PCR analysis showed that the two genes together constitute an operon. NCgl2816 putatively codes for a lactate permease, while NCgl2817 was demonstrated to encode quinone-dependent l-lactate dehydrogenase, which was named LldD. C. glutamicum LldD displayed Michaelis-Menten kinetics for the substrate l-lactate with a K(m) of about 0.51 mM. The specific activity of LldD was about 10-fold higher during growth on l-lactate or on an l-lactate-glucose mixture than during growth on glucose, d-lactate, or pyruvate, while the specific activity of quinone-dependent d-lactate dehydrogenase differed little with the carbon source. RNA levels of NCgl2816 and lldD were about 18-fold higher during growth on l-lactate than on pyruvate. Disruption of the NCgl2816-lldD operon resulted in loss of the ability to utilize l-lactate as the sole carbon source. Expression of lldD restored l-lactate utilization, indicating that the function of the permease gene NCgl2816 is dispensable, while LldD is essential, for growth of C. glutamicum on l-lactate.

Bacterial Proteins↗

PubMed2004

LuxS is required for persistent pneumococcal carriage and expression of virulence and biosynthesis genes.

Streptococcus pneumoniae causes several diseases, including otitis media, pneumonia, and meningitis. Although little is known about the regulation of or how individual pneumococcal factors contribute to these disease states, there is evidence suggesting that some factors are regulated by a cell-density-dependent mechanism (quorum sensing). Quorum sensing allows bacteria to couple transcription with changes in cell density; bacteria achieve this by sensing and responding to small diffusible signaling molecules. We investigated how the LuxS signaling system impacts the biology of S. pneumoniae. An analysis of the transcriptional profiles of a serotype 2 strain and an isogenic luxS deletion strain utilizing an S. pneumoniae-specific microarray indicated that LuxS regulates gene expression involved in discrete cellular processes, including pneumolysin expression. Contrary to the paradigm for quorum sensing, we observed pronounced effects on transcription in early log phase, where gene expression was repressed in the mutant. Assessing the mutant for its ability to infect and cause disease in animals revealed a profound defect in ability to persist in the nasopharyngeal tissues. Our analysis of an S. pneumoniae transcriptome revealed a function for LuxS in gene regulation that is not dependent upon high cell density and is likely involved in the maintenance of pneumococcal load in susceptible hosts.

Bacterial Proteins↗

PubMed2026

De Novo TRIO Missense Variants Disrupt Ras-GEF Domains and Cause Congenital Ventriculomegaly and Hydrocephalus.

Congenital hydrocephalus (CH), characterized by congenital ventriculomegaly (CV), affects approximately 0.5-1 per 1000 live births and is a common cause of pediatric neurosurgical intervention, yet its genetic architecture remains incompletely defined. We report a child with syndromic CH requiring cerebrospinal fluid diversion who harbored a pathogenic de novo missense variant in TRIO (c.3232C > T; p.(Arg1078Trp)), a gene previously associated with autosomal dominant neurodevelopmental disorders featuring variable head circumference. This case prompted systematic evaluation of TRIO variation in our CV/CH cohort (2,697 patient-parent trios) using exome sequencing. We identified five additional unrelated probands with de novo TRIO variants, including two novel substitutions affecting the same residue within the Ras-GEF1 domain (p.(Glu1299Lys) and p.(Glu1299Gly)), yielding significant gene-level enrichment for protein-damaging de novo variants (adjusted p = 6.12 × 10-5). All affected individuals exhibited CV, frequently accompanied by developmental delay and additional structural brain abnormalities. In silico structural modeling predicted that associated variants destabilize critical TRIO Ras-GEF domains required for Rho GTPase activation. Analysis of single-nucleus transcriptomic data from the developing human neocortex revealed enrichment of TRIO expression in multipotent progenitor populations. A systematic literature review identified six additional individuals with TRIO de novo variants and reported CV or CH, including an unrelated patient with the same p.(Arg1078Trp) substitution. Together, these findings expand the phenotypic spectrum associated with pathogenic TRIO variation to include CV/CH and support TRIO as a clinically relevant gene in the genetic evaluation of syndromic CV/CH patients.

Humans↗

PubMed2026

Integrative Multiomics and Drug Sensitivity Profiling Reveal Potential Biomarkers and Therapeutic Strategies in Pediatric Solid Tumors.

UNLABELLED: Cure rates for childhood malignancies using established therapy protocols have increased to an average of 80% but have reached a plateau. Moreover, survival rates are particularly low for some pediatric tumors-such as high-risk group 3 medulloblastomas, osteosarcomas, Ewing sarcomas, high-risk neuroblastomas, and high-grade gliomas-and dismal for patients with relapsed malignancies. A functional drug response profiling platform for pediatric solid and brain tumors has been established within the INFORM program to identify patient-specific vulnerabilities and biomarkers and to unravel molecular mechanisms associated with drug response profiles for clinical translation. In this study, we performed a multiomics analysis using drug sensitivity profiles, as well as genomic and transcriptomic data, of 81 pediatric solid tumor samples. The integrative analysis suggested two multiomics signatures associated with drug sensitivity. One signature distinguished neuroblastoma samples with sensitivity to navitoclax, a BCL2 family inhibitor. A second signature was specific to a subset of Wilms tumors harboring the SIX1 (Q177R) hotspot mutation that displayed high expression of MGAM, PTPN14, STAT4, and KDM2B and high sensitivity to MEK inhibitors. A patient-specific causal interaction network analysis suggested possible molecular interactions between MEK inhibitors and the SIX1 mutation in Wilms tumor samples. In conclusion, the integration of drug sensitivity profiling and multiomics data revealed potential biomarkers that may be associated with drug sensitivity in pediatric solid tumors. Patient-specific causal interaction network analysis further elucidated the interaction between inhibitors and signature biomarkers, providing insights that may inform clinical translation. SIGNIFICANCE: The combination of multiomics analysis and drug sensitivity profiling identified two signatures related to drug sensitivity in pediatric solid tumors, contributing to the advancement of functional precision medicine and personalized treatment strategies. This article is part of a special series: Driving Cancer Discoveries with Computational Research, Data Science, and Machine Learning/AI .

Humans↗

PubMed2002

Comparative mycobacterial genomics as a tool for drug target and antigen discovery.

Genomics and the associated downstream technologies are generating vast data sets that provide new opportunities for understanding and combating both infectious and genetic diseases in humans. The genomic approach has been applied to tuberculosis, a major cause of transmissible morbidity and mortality, with notable success. Complete genome sequences are now available for three members of the Mycobacterium tuberculosis complex and the related intracellular pathogen M. leprae. Many of the predictions generated in silico by genomics have been validated through functional analysis, including studies of the transcriptome and proteome, and led to the identification of essential genes. Knowledge of the latter defines potential targets for new and existing drugs and their specificity can be assessed by comparative genomics with the host or other pathogens. Genomics is also furthering tuberculosis vaccine development by pinpointing potentially antigenic proteins as well as providing better diagnostic tools to detect infection.

Antigens↗

PubMed2026

Downregulated lysyl oxidase in plasma extracellular vesicles: a biomarker linked to brain metastasis risk in lung adenocarcinoma.

BACKGROUND: Brain metastasis (BrM) is a leading cause of mortality in patients with lung adenocarcinoma (LUAD). Extracellular vesicles (EVs), which carry bioactive molecules, play a critical role in tumor microenvironment remodeling and exhibit metastatic organotropism, holding promise as liquid biopsy biomarkers. This study aims to identify plasma EV-derived proteins associated with LUAD-BrM. METHODS: A multi-omics framework was applied. Plasma EVs from 59 stage IV LUAD patients (30 BrM vs 29 non-BrM) were profiled using data-independent acquisition mass spectrometry proteomics. Candidate proteins were screened via bioinformatics and machine learning (LASSO/RF/SVM). Initial validation included tissue proteomics (n = 13), single-cell transcriptomics (TISCH2), and Western blot analysis of a subset of the discovery samples. Functional experiments were conducted in vitro. The lead candidate was ultimately validated in an independent plasma cohort (n = 158) through ELISA. RESULTS: Proteomic analysis implicated collagen-containing extracellular matrix (ECM) pathways. Lysyl oxidase (LOX), a key ECM cross-linking enzyme, was identified as a lead candidate. LOX and its family member LOXL1 were consistently downregulated in BrM tissues and plasma EVs. Single-cell analysis revealed decreased LOX expression specifically in BrM-associated fibroblasts, which showed suppressed ECM-related pathways. In vitro experiments supported a PI3K/AKT-LOX-ECM regulatory axis. Plasma EV-derived LOX demonstrated strong diagnostic performance in the independent cohort, with an AUC of 0.786 (95% CI 0.713iated fi. CONCLUSIONS: Our study establishes plasma EV-derived LOX as a promising non-invasive biomarker for LUAD-BrM through a comprehensive multi-omics validation strategy. We propose a model wherein downregulation of LOX, potentially driven by PI3K/AKT signaling in tumor-associated fibroblasts, contributes to ECM degradation and may promote brain-tropic metastasis. This finding offers new insights for risk stratification and timely intervention in LUAD patients.

Humans↗

PubMed2025

CAFs activated by YAP1 upregulate cancer matrix stiffness to mediate hepatocellular carcinoma progression.

BACKGROUND: The stiffness of the matrix is closely related to the progression of hepatocellular carcinoma (HCC). Although direct targeting of stromal rigidity in HCC remains a clinical challenge, cancer-associated fibroblasts (CAFs) are considered key contributors to this process. Given the heterogeneity of CAFs, this study explored the relationship between specific CAF subsets and liver cancer matrix stiffness, aiming to identify novel therapeutic targets for HCC patients. METHODS: Single-cell sequencing datasets were leveraged to identify cell types within liver cancer and characterize the transcriptomic profiles of CAFs. Prognostic analysis, utilizing the Gene Expression Profiling Interactive Analysis (GEPIA) and The Cancer Genome Atlas (TCGA) liver cancer datasets, assessed the correlation between matrix stiffness-related genes and HCC patient outcomes. Pseudo-time analysis was applied to trace the developmental trajectories of CAFs. By calculating intercellular communication probabilities and analyzing transcription factor activity, the functions and interactions of different CAF subsets were elucidated. Gene Ontology (GO) analysis was used to explore the functional roles of CAFs in distinct Yes-associated protein 1 (YAP1) groups. Finally, cellular experiments and animal experiments were further conducted to validate the hypotheses of this study. RESULTS: This study identified CAF subpopulations based on single-cell sequencing data and analyzed transcriptional changes within these subpopulations. Key findings include the identification of collagen type I alpha 1 (COL1A1), collagen type III alpha 1 (COL3A1), and lysyloxidase (LOX) as pivotal node genes during CAF development. Moreover, the expression of matrix stiffness-related genes was inversely correlated with the prognosis of HCC patients. Notably, the YAP1-positive CAF subpopulation emerged as the primary contributor to matrix stiffness in liver cancer. This subpopulation upregulates the expression of matrix stiffness-related genes and promotes tumor progression by activating signaling pathways such as autophagy and GTPase activity regulation. Cellular experiments and animal studies further validated this conclusion. CONCLUSION: This single-cell analysis uncovered the functional roles of CAFs in liver cancer. The YAP1-positive CAF subpopulation, in particular, was shown to contribute to matrix stiffness by upregulating the expression of relevant genes and promoting tumor progression through the activation of specific signaling pathways.

Carcinoma, Hepatocellular↗

PubMed2006

A novel secreted endonuclease from Culex quinquefasciatus salivary glands.

Previous analysis of the salivary gland transcriptome of Culex quinquefasciatus showed the potential presence of an endonuclease with sequence similarities to shrimp, crab and two tsetse salivary proteins. Indeed, not only was the cloned cDNA shown to encode an active double-stranded endonuclease, but also the same activity was demonstrated to be secreted by salivary glands of Cx. quinquefasciatus. Preliminary studies with salivary gland extracts confirmed the presence of a highly active nuclease. This enzyme was shown to be present in the saliva of female mosquitoes by allowing starved mosquitoes to probe DNA-containing agarose gel. The recombinant Cx. quinquefasciatus endonuclease (CuquEndo) produced in mammalian cells showed no sequence specificity for DNA substrate except that it only cleaves double-stranded DNA. Recombinant Cx. quinquefasciatus endonuclease was active in the presence of Mg(2+) ions at pH 7.0-8.0, but no endonuclease activity was detected in the presence of calcium ions. The final hydrolysis products of this enzyme, detected by ion exchange chromatography, yielded DNA fragments ranging form 8-12 base pairs. Although endonucleases have been associated with a variety of cellular functions, their role in mosquito saliva is not clear. This female-specific secreted endonuclease may assist blood meal intake by lowering the local viscosity created by the release of host DNA in the bite site and/or acting as an indirect anticoagulant factor by producing a defibrotide-like mixture of DNA haptamers.

Amino Acid Sequence↗

PubMed2025

Proper 5'-3' cotranslational mRNA decay in yeast requires import of Xrn1 to the nucleus.

The budding yeast Xrn1 protein shuttles between the nucleus, where it stimulates transcription, and the cytoplasm, where it executes the major cytoplasmic mRNA decay. In the cytoplasm, apart from catalyzing 5'→3' decay onto non translated mRNAs, Xrn1 can follow the last translating ribosome to degrade the decapped mRNA template, a process known as "cotranslational mRNA decay". We have previously observed that the import of Xrn1 to the nucleus is required for efficient cytoplasmic mRNA decay. Here by using an Xrn1 mutant that cannot enter the nucleus, but is otherwise functional in ribonuclease activity, we show that nuclear import is necessary for proper global cotranslational decay of mRNAs along coding regions and also affects degradation in the of 5' region of a large group of mRNAs, which comprise about 20% of the transcriptome. Furthermore, a principal component analysis of the genomic datasets of this mutant and other Xrn1 mutants also shows that lack of a cytoplasmic 5'→3' exoribonuclease is the primary cause of the physiological defects seen in a xrn1Δ mutant, but also suggests that Xrn1 import into the nucleus is necessary for its full in vivo functions.

Exoribonucleases↗

PubMed2005

Differential gene expression profiling of human umbilical cord blood-derived mesenchymal stem cells by DNA microarray.

Mesenchymal stem cells (MSCs) retain both self-renewal and multilineage differentiation capabilities. Despite wide therapeutic potential, many aspects of human MSCs, particularly the molecular parameters to define the stemness, remain largely unknown. Using high-density oligonucleotide micro-arrays, we obtained the differential gene expression profile between a fraction of mononuclear cells of human umbilical cord blood (UCB) and its MSC subpopulation. Of particular interest was a subset of 47 genes preferentially expressed at 50-fold or higher in MSCs, which could be regarded as a molecular foundation of human MSCs. This subset contains numerous genes encoding collagens, other extracellular matrix or related proteins, cytokines or growth factors, and cytoskeleton-associated proteins but very few genes for membrane and nuclear proteins. In addition, a direct comparison of this microarray-generated transcriptome with the published serial analysis of gene expression data suggests that a molecular context of UCB-derived MSCs is more or less similar to that of bone marrow-derived cells. Altogether, our results will provide a basis for studies on molecular mechanisms controlling core properties of human MSCs.

Bone Marrow Cells↗

PubMed2002

Proteomic approaches to central nervous system disorders.

The discovery, design and evaluation of new medicines is critically dependent on the elucidation of protein mechanisms involved in human diseases. Since the proteome of a cell or tissue is not a simple reflection of its transcriptome, direct protein-based analysis is needed. Advances in proteomic technologies are improving the analysis of membrane proteins and signaling complexes with increased speed and molecular detail. Changes in protein isoforms due to post-translational modifications, such as phosphorylation induced by cell signaling events and alternative splice forms of receptors, may be mapped to an altered protein expression pattern in clinically relevant cell populations with a causative or diagnostic disease link. A CNS proteome database derived from primary human tissues may avoid ambiguities of experimental models. It will also accelerate the development of more specific diagnostic and prognostic disease markers as well as new selective therapeutics. Proteomics is also being applied to resolve in silico gene prediction uncertainties by direct open reading frame verification. These advances hold great promise for improvements in the understanding, diagnosis and therapy of central nervous system disorders.

Biomarkers↗

PubMed2001

Transcriptome of a mouse kidney cortical collecting duct cell line: effects of aldosterone and vasopressin.

Aldosterone and vasopressin are responsible for the final adjustment of sodium and water reabsorption in the kidney. In principal cells of the kidney cortical collecting duct (CCD), the integral response to aldosterone and the long-term functional effects of vasopressin depend on transcription. In this study, we analyzed the transcriptome of a highly differentiated mouse clonal CCD principal cell line (mpkCCD(cl4)) and the changes in the transcriptome induced by aldosterone and vasopressin. Serial analysis of gene expression (SAGE) was performed on untreated cells and on cells treated with either aldosterone or vasopressin for 4 h. The transcriptomes in these three experimental conditions were determined by sequencing 169,721 transcript tags from the corresponding SAGE libraries. Limiting the analysis to tags that occurred twice or more in the data set, 14,654 different transcripts were identified, 3,642 of which do not match known mouse sequences. Statistical comparison (at P < 0.05 level) of the three SAGE libraries revealed 34 AITs (aldosterone-induced transcripts), 29 ARTs (aldosterone-repressed transcripts), 48 VITs (vasopressin-induced transcripts) and 11 VRTs (vasopressin-repressed transcripts). A selection of the differentially-expressed, hormone-specific transcripts (5 VITs, 2 AITs and 1 ART) has been validated in the mpkCCD(cl4) cell line either by Northern blot hybridization or reverse transcription-PCR. The hepatocyte nuclear transcription factor HNF-3-alpha (VIT39), the receptor activity modifying protein RAMP3 (VIT48), and the glucocorticoid-induced leucine zipper protein (GILZ) (AIT28) are candidate proteins playing a role in physiological responses of this cell line to vasopressin and aldosterone.

Aldosterone↗

PubMed2026

Single-cell transcriptomics reveals that air-liquid interface culture promotes goblet cell differentiation and inhibits glycolysis in organoid cell monolayers.

Faithfully recapitulating the cellular heterogeneity of the intestinal epithelium is essential when using organoid models. Air-liquid interface (ALI) culture has been shown to promote secretory cell differentiation, but its impact on gene expression in each epithelial cell type remains unclear. In this study, we used single-cell RNA sequencing (scRNA-seq) to characterize the cellular heterogeneity of rabbit cecum-derived organoid monolayers grown under immerged or ALI conditions. We then compared these organoid cell type-specific gene expression profiles to a scRNA-seq atlas of the rabbit cecal epithelium in vivo. We selected the rabbit model notably because, unlike mice, it possesses BEST4+ epithelial cells, a newly discovered subset of mature absorptive cells. Our analysis revealed a high degree of transcriptomic similarity between in vivo and organoid-derived stem and transit-amplifying cells. ALI culture markedly enhanced the differentiation of the secretory lineage, especially goblet cells, whose transcriptome closely resembled that of in vivo goblet cells. Furthermore, ALI was the only condition allowing the detection of enteroendocrine cells. BEST4+ cells, however, were absent from organoids in immerged or ALI conditions despite their presence in vivo. In addition, ALI culture led to a consistent downregulation of hypoxia and glycolysis-associated genes across all cell types, which suggests a metabolic shift likely driven by increased oxygen availability in ALI conditions. Cell-cell communication analyses further indicated that ALI more closely mirrored in vivo patterns than immerged condition. Altogether, these results demonstrate that ALI culture allows for better recapitulation of the in vivo cellular heterogeneity and molecular signatures of the intestinal epithelium.NEW & NOTEWORTHY Using single-cell RNA sequencing, this study shows that air-liquid interface (ALI) culture enhances secretory lineage differentiation of intestinal organoid cell monolayers and improves transcriptomic similarity to the native epithelium. ALI reduced hypoxia-associated gene expression and better recapitulates in vivo-like cell-cell interactions, supporting its value for modeling intestinal epithelial heterogeneity in organoids.

Animals↗

PubMed2026

Beyond ion channel dysfunction: Integration of the transcriptome and proteome from patient-specific re-engineered cardiac cells, and population-level QT genome-wide association study reveals broad cellular dysfunction.

BACKGROUND: Congenital long QT syndrome (LQTS) is a cardiac channelopathy with increased risk of cardiac-triggered syncope/seizures, sudden cardiac arrest, and sudden cardiac death. OBJECTIVE: This study aimed to describe the transcriptomic and proteomic profiles in patient-derived inducible pluripotent stem cell-derived cardiomyocyte (iPSC-CM) models of the 3 canonical genotypes of congenital LQTS: LQT1, LQT2, and LQT3 and integrate these omics-level findings with each other and with population/clinical level QT-genome-wide association study (GWAS) data. METHODS: LQT1, LQT2, LQT3 and respective isogenic control iPSC-CMs were cultured, and RNA and protein samples were collected. RNA sequencing and mass spectrometry-enabled proteomic analysis was performed. PrediXcan analysis was performed using QT GWAS summary statistics and transcriptome expression data. Differential gene and protein expression and ingenuity pathway analysis (IPA) was performed comparing each LQT genotype with its respective isogenic control. RESULTS: 1645 differentially expressed genes (DEGs) were identified; 13 were altered in all 3 LQTS genotypes. IPA analysis of DEGs revealed 301 altered pathways; 47 were altered in all LQTS genotypes. Proteomic analysis identified 2561 differentially expressed proteins (DEPs); 30 were altered in all 3 genotypes. IPA analysis of DEPs identified 646 altered pathways. 306 genes/proteins were identified as significantly altered in both the transcriptome and proteome; pathway analysis of these 301 genes identified 201 altered pathways. 7 pathways were altered in all 3 LQTS genotypes in both the transcriptome and proteome. Integration of the population-level PrediXcan results and the cardiomyocyte-derived omics results identified multiple shared pathways. CONCLUSION: Multi-omics analysis of LQTS and integration of omics results with QT GWAS data reveals that primary LQTS-causative ion channel defects precipitate secondary alterations in a wide range of cellular pathways. Our findings suggest more broad molecular level changes throughout the cell. This study lays the foundation for further exploration of broad cellular changes resulting from ion channel disturbances and how they contribute to disease mechanism.

Humans↗