Cell Type-specific Expression Research Articles

Root cell-type specific expressions of bacterial mercury transporter MerC and plant SNARE SYP121 fusion protein differentially affect cadmium accumulation patterns of Arabidopsis

ABSTRACT There is increasing demand for solutions against cadmium pollution to secure food safety, and phytoremediation is one of the potential tools. We previously found that a bacterial mercury transporter MerC possesses cadmium uptake activity and its overexpression as a fusion protein with a plasma-membrane resident SNARE protein, SYP121 enhances the cadmium uptake ability of Arabidopsis plants. In this study, we examined whether two different root cell-type specific expression systems of MerC-SYP121 fusion protein could efficiently enhance cadmium accumulation of Arabidopsis plants, compared to the p35S-driven ubiquitous expression system. Representative transgenic lines expressing MerC-SYP121 in root surface cells (pEpi lines) or root endodermal cells (pSCR lines), established in our previous studies, were subjected to different cadmium treatments along with the p35S line. A vertical agar plate assay showed that root surface-specific line pEpi, as well as the p35S line, showed about 15% higher cadmium accumulation in shoots after one-day 10 µM cadmium treatment, compared to the wild-type Col-0. On the other hand, the endodermis-specific line pSCR accumulated 30% less cadmium in its shoots. A similar cadmium accumulation pattern in shoots was observed under the environmentally relevant much lower cadmium treatment of 0.1 µM for 4 d, using the hydroponic culture system. To further examine the potential of the MerC-SYP121 expression system for cadmium phytoremediation, the transgenic plants were hydroponically exposed to 0.1 µM for 4 weeks. The cadmium accumulation after the 4 weeks of treatment was again 16% higher in the pEpi shoots compared to that of Col-0, whereas the p35S line only showed 6% higher Cd concentration. Shoots of the pSCR line accumulated slightly less cadmium compared to Col-0. Ionomic profiles in these plants were analyzed, however, pSCR-specific patterns were not evident. Nevertheless, in our previous studies, pEpi and pSCR lines both efficiently accumulated more mercury in shoots than in the wild-type. The presented results suggest that the effects of cell-type-specific MerC-SYP121 expression differ by the target metals and its expression in root surface cells rather than that in endodermis is suitable for enhancing root cadmium uptake and subsequent shoot accumulation.

Genetics of SLE: mechanistic insights from monogenic disease and disease-associated variants.

The past few years have provided important insights into the genetic architecture of systemic autoimmunity through aggregation of findings from genome-wide association studies (GWAS) and whole-exome or whole-genome sequencing studies. In the prototypic systemic autoimmune disease systemic lupus erythematosus (SLE), monogenic disease accounts for a small fraction of cases but has been instrumental in the elucidation of disease mechanisms. Defects in theclearance or digestion of extracellular or intracellular DNA or RNA lead to increased sensing of nucleic acids, which can break B cell tolerance and induce the production of type I interferons leading to tissue damage. Current data suggest that multiple GWAS SLE risk alleles act in concert with rare functional variants to promote SLE development. Moreover, introduction of orthologous variant alleles into mice has revealed that pathogenic X-linked dominant and recessive SLE can be caused by novel variants in TLR7 and SAT1, respectively. Such bespoke models of disease help tounravel pathogenic pathways and can be used to test targeted therapies. Cell type-specific expression data revealed that most GWAS SLE risk genes are highly expressed in age-associated B cells (ABCs), which supports the view that ABCs produce lupus autoantibodies and contribute to end-organ damage by persisting in inflamed tissues, including the kidneys. ABCs have thus emerged as key targets of promising precision therapeutics.

Identification of key genes and the pathophysiology associated with allergen-specific immunotherapy for allergic rhinitis

BackgroundAllergen-specific immunotherapy (AIT) is a causative treatment in allergic rhinitis (AR), comprising long-term allergen administration and over three years of treatment. This study is carried out for revealing the mechanisms and key genes of AIT in AR.MethodsThe present study utilized online Gene Expression Omnibus (GEO) microarray expression profiling dataset GSE37157 and GSE29521 to analyze the hub genes changes related to AIT in AR. Based on limma package, differential expression analysis for the two groups (samples of allergic patients prior to AIT and samples of allergic patients undergoing AIT) was performed to obtain differentially expressed genes (DEGs). Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs were conducted using DAVID database. A Protein-Protein Interaction network (PPI) was built and a significant network module was acquired by using Cytoscape software (Cytoscape, 3.7.2). Utilizing the miRWalk database, we identified potential gene biomarkers, constructed interaction networks of target genes and microRNAs (miRNAs) using Cytoscape software, and explore the cell type-specific expression patterns of these genes in peripheral blood using publicly available single-cell RNA sequencing data (GSE200107). Finally, we are using PCR to detect changes in the hub genes that are screened using the above method in peripheral blood before and after AIT treatment.ResultsGSE37157 and GSE29521 included 28 and 13 samples, respectively. A total of 119 significantly co-upregulated DEGs and 33 co-downregulated DEGs were obtained from two datasets. The GO and KEGG analyses demonstrated that protein transport, positive regulation of apoptotic process, Natural killer cell mediated cytotoxicity, T cell receptor signaling pathway, TNF signaling pathway, B cell receptor signaling pathway and Apoptosis may be potential candidate therapeutic targets for AIT of AR. From the PPI network, 20 hub genes were obtained. Among them, the PPI sub-networks of CASP3, FOXO3, PIK3R1, PIK3R3, ATF4, and POLD3 screened out from our study have been identified as reliable predictors of AIT in AR, especially the PIK3R1.ConclusionOur analysis has identified novel gene signatures, thereby contributing to a more comprehensive understanding of the molecular mechanisms underlying AIT in the treatment of AR.

Cross-Tissue Single-Nucleus RNA Sequencing Discovers Tissue-Resident Adipocytes Involved in Propanoate Metabolism in the Human Heart.

Adipocytes are crucial regulators of cardiovascular health. However, not much is known about gene expression profiles of adipocytes residing in nonfat cardiovascular tissues, their genetic regulation, and contribution to coronary artery disease. Here, we investigated whether and how the gene expression profiles of adipocytes in the subcutaneous adipose tissue differ from adipocytes residing in the heart. We used single-nucleus RNA-sequencing data sets of subcutaneous adipose tissue and heart and performed in-depth analysis of tissue-resident adipocytes and their cell-cell interactions. We first discovered tissue-specific features of tissue-resident adipocytes, identified functional pathways involved in their tissue specificity, and found genes with cell type-specific expression enrichment in tissue-resident adipocytes. By following up these results, we discovered the propanoate metabolism pathway as a novel distinct characteristic of the heart-resident adipocytes and found a significant enrichment of coronary artery disease genome-wide association study risk variants among the right atrium-specific adipocyte marker genes. Our cell-cell communication analysis identified 22 specific heart adipocyte-associated ligand-receptor pairs and signaling pathways, including THBS (thrombospondin) and EPHA (ephrin type-A), further supporting the distinct tissue-resident role of heart adipocytes. Our results also suggest chamber-level coordination of heart adipocyte expression profiles as we observed a consistently larger number of adipocyte-associated ligand-receptor interactions and functional pathways in the atriums than ventricles. Overall, we introduce a new function and genetic link to coronary artery disease for the previously unexplored heart-resident adipocytes.

Cell type-specific expression of the molecular players in mouse prefrontal cortex during cocaine addiction

Cocaine addiction is an issue that affects more than 5 million people in America per year. Although there has been much research into the genes and chemicals responsible for cocaine addiction, there are many specific questions left unanswered. Our experiment attempts to further previous research into certain molecular players. We follow up with their use of single-cell RNA sequencing on the prefrontal cortex cells of mice undergoing cocaine intravenous self-administration. Data of 12 samples from both saline and cocaine treated mice which are found on the Gene Expression Omnibus public database were retrieved. Using the Seurat function of RStudio, the data was merged into objects, normalized, clustered, and labeled into one of eight cell types. What resulted was a detailed UMAP plot displaying the clusters, their gene expression level, expression frequency, and their cell type. With this plot, we were able to determine the specific cell types that express the genes encoding the pre-established molecular players (∆FosB, MeCP2, and BDNF). When the analysis was expanded to a cell-type specific level, it was discovered some of these genes were selectively expressed in excitatory neurons and non-neuronal cells. Going further into the analysis, we determined the 6 genes with the most varied gene expression over the 3 stages of cocaine addiction for each of the 8 cell types. Overall, our computational analysis of publicly available transcriptome datasets from mouse addiction model provides new insights into the molecular basis of cocaine addiction.

Cell Type-Specific Promoters of Volvox carteri for Molecular Cell Biology Studies.

The multicellular green alga Volvox carteri has emerged as a valuable model organism for investigating various aspects of multicellularity and cellular differentiation, photoreception and phototaxis, cell division, biogenesis of the extracellular matrix and morphogenetic movements. While a range of molecular tools and bioinformatics resources have been made available for exploring these topics, the establishment of cell type-specific promoters in V. carteri has not been achieved so far. Therefore, here, we conducted a thorough screening of transcriptome data from RNA sequencing analyses of V. carteri in order to identify potential cell type-specific promoters. Eventually, we chose two putative strong and cell type-specific promoters, with one exhibiting specific expression in reproductive cells (gonidia), the PCY1 promoter, and the other in somatic cells, the PFP promoter. After cloning both promoter regions, they were introduced upstream of a luciferase reporter gene. By using particle bombardment, the DNA constructs were stably integrated into the genome of V. carteri. The results of the expression analyses, which were conducted at both the transcript and protein levels, demonstrated that the two promoters drive cell type-specific expression in their respective target cell types. Transformants with considerably diverse expression levels of the chimeric genes were identifiable. In conclusion, the screening and analysis of transcriptome data from RNA sequencing allowed for the identification of potential cell type-specific promoters in V. carteri. Reporter gene constructs demonstrated the actual usability of two promoters. The investigated PCY1 and PFP promoters were proven to be potent molecular tools for genetic engineering in V. carteri.

Downregulation of CCR5 on brain perivascular macrophages in simian immunodeficiency virus-infected rhesus macaques.

C-C chemokine receptor 5 (CCR5) is a major coreceptor for Human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) cell entry; however, its role in brain pathogenesis is largely understudied. Thus, we sought to examine cell type-specific protein expression of CCR5 during SIV infection of the brain. We examined occipital cortical tissue from uninfected rhesus macaques and SIV-infected animals with or without encephalitis using immunohistochemistry and immunofluorescence microscopy to determine the number and distribution of CCR5-positive cells. An increase in the number of CCR5+ cells in the brain of SIV-infected animals with encephalitis was accounted for by increased CD3+CD8+ cells expressing CCR5, but not by increased CCR5+ microglia or perivascular macrophages (PVMs), and a concurrent decrease in the percentage of CCR5+ PVMs was observed. Levels of CCR5 and SIV Gag p28 protein expression were examined on a per-cell basis, and a significant, negative relationship was established indicating decreased CCR5 expression in productively infected cells. While investigating the endocytosis-mediated CCR5 internalization as a mechanism for CCR5 downregulation, we found that phospho-ERK1/2, an indicator of clathrin-mediated endocytosis, was colocalized with infected PVMs and that macrophages from infected animals showed significantly increased expression of clathrin heavy chain 1. These findings show a shift in CCR5-positive cell types in the brain during SIV pathogenesis with an increase in the number of CCR5+ CD8 T cells, and downregulated CCR5 expression on infected PVMs, likely through ERK1/2-driven, clathrin-mediated endocytosis.

WAPL functions as a rheostat of Protocadherin isoform diversity that controls neural wiring.

Neural type-specific expression of clustered Protocadherin (Pcdh) proteins is essential for the establishment of connectivity patterns during brain development. In mammals, deterministic expression of the same Pcdh isoform promotes minimal overlap of tiled projections of serotonergic neuron axons throughout the brain, while stochastic expression of Pcdh genes allows for convergence of tightly packed, overlapping olfactory sensory neuron axons into targeted structures. How can the same gene locus generate opposite transcriptional programs that orchestrate distinct spatial arrangements of axonal patterns? Here, we reveal that cell type-specific Pcdh expression and axonal behavior depend on the activity of cohesin and its unloader, WAPL (wings apart-like protein homolog). While cohesin erases genomic-distance biases in Pcdh choice, WAPL functions as a rheostat of cohesin processivity that determines Pcdh isoform diversity.

Transcription factors KANADI 1, MYB DOMAIN PROTEIN 44, and PHYTOCHROME INTERACTING FACTOR 4 regulate long intergenic noncoding RNAs expressed in Arabidopsis roots.

Thousands of long intergenic noncoding RNAs (lincRNAs) have been identified in plant genomes. While some lincRNAs have been characterized as important regulators in different biological processes, little is known about the transcriptional regulation for most plant lincRNAs. Through the integration of 8 annotation resources, we defined 6,599 high-confidence lincRNA loci in Arabidopsis (Arabidopsis thaliana). For lincRNAs belonging to different evolutionary age categories, we identified major differences in sequence and chromatin features, as well as in the level of conservation and purifying selection acting during evolution. Spatiotemporal gene expression profiles combined with transcription factor (TF) chromatin immunoprecipitation (ChIP) data were used to construct a TF-lincRNA regulatory network containing 2,659 lincRNAs and 15,686 interactions. We found that properties characterizing lincRNA expression, conservation, and regulation differ between plants and animals. Experimental validation confirmed the role of 3 TFs, KANADI 1, MYB DOMAIN PROTEIN 44, and PHYTOCHROME INTERACTING FACTOR 4, as key regulators controlling root-specific lincRNA expression, demonstrating the predictive power of our network. Furthermore, we identified 58 lincRNAs, regulated by these TFs, showing strong root cell type-specific expression or chromatin accessibility, which are linked with genome-wide association studies genetic associations related to root system development and growth. The multilevel genome-wide characterization covering chromatin state information, promoter conservation, and chromatin immunoprecipitation-based TF binding, for all detectable lincRNAs across 769 expression samples, permits rapidly defining the biological context and relevance of Arabidopsis lincRNAs through regulatory networks.

69-LB: Sex-Specific Dynamics of GLP-1R and GIPR Expression in the Hypothalamus and the Amygdala of Mice

GIPR:GLP-1R co-agonism improves obesity and cardio-metabolic disease with superior efficacy in both male and female mice compared to mono-agonism. We detected sex-specific differences in metabolic phenotypes, which may point to differing sensitivity for GLP-1R and/or GIPR in areas of the hypothalamus regulating energy balance (ARC, VMH, DMH) and in areas of the amygdala regulating cardiovascular functions. To optimize future therapeutic benefits and minimize side effects from a perspective of sex differences, we investigated whether GLP1R and GIPR expression levels in specific cell populations of the hypothalamus and the amygdala regions are co-determined by sex and diet. Female and male C57Bl6 mice were fed normal chow or 58% high fat diet (HFD) for 21 days (n = 4, total mouse number 16). Spatial transcriptomics by in-situ hybridization method RNAscope™ was combined with immunofluorescence staining for neurons (NeuN), astrocytes (GFAP) and microglia (iba1) to identify cell type specific gene expression patterns of GLP1R and GIPR in female and male brains. In the hypothalamus, both sexes, gene expression of GLP-1R was higher compared to GIPR (ARC, VMH and DMH). In our studies GLP-1R and GIPR were mostly expressed in neurons, marginally in microglia and not observed in astrocytes in both sexes. HFD exposure reduced GLP-1R and GIPR gene expression in males only. In the Amygdala, neuronal GIPR gene expression levels were higher compared to GLP-1R in both sexes. HFD reduced GIPR and GLP-1R gene expression in males only. In females, HFD induced a slight increase of GIPR gene expression specifically in the central and basolateral amygdala. Our results reveal sex-specific dynamics in GLP-1R and GIPR expression patterns in metabolically relevant areas of the hypothalamus and the amygdala induced by HFD exposure. This may imply sex-specific dosing regimens of GIP:GLP-1R co-agonists for future treatments of cardio-metabolic disease Disclosure S. M. Hofmann: None. S. Zhang: None. R. Tom: None. T. D. Müller: Research Support; Novo Nordisk A/S, Speaker's Bureau; Eli Lilly and Company, AstraZeneca, Mercodia AB, Stock/Shareholder; Novo Nordisk A/S, Eli Lilly and Company. M. H. Tschöp: Consultant; Boehringer Ingelheim Pharma GmbH&Co. KG.

RELA governs a network of islet-specific metabolic genes necessary for beta cell function

Aims/hypothesisNF-κB activation unites metabolic and inflammatory responses in many diseases yet less is known about the role that NF-κB plays in normal metabolism. In this study we investigated how RELA impacts the beta cell transcriptional landscape and provides network control over glucoregulation.MethodsWe generated novel mouse lines harbouring beta cell-specific deletion of either the Rela gene, encoding the canonical NF-κB transcription factor p65 (βp65KO mice), or the Ikbkg gene, encoding the NF-κB essential modulator NEMO (βNEMOKO mice), as well as βA20Tg mice that carry beta cell-specific and forced transgenic expression of the NF-κB-negative regulator gene Tnfaip3, which encodes the A20 protein. Mouse studies were complemented by bioinformatics analysis of human islet chromatin accessibility (assay for transposase-accessible chromatin with sequencing [ATAC-seq]), promoter capture Hi-C (pcHi-C) and p65 binding (chromatin immunoprecipitation–sequencing [ChIP-seq]) data to investigate genome-wide control of the human beta cell metabolic programme.ResultsRela deficiency resulted in complete loss of stimulus-dependent inflammatory gene upregulation, consistent with its known role in governing inflammation. However, Rela deletion also rendered mice glucose intolerant because of functional loss of insulin secretion. Glucose intolerance was intrinsic to beta cells as βp65KO islets failed to secrete insulin ex vivo in response to a glucose challenge and were unable to restore metabolic control when transplanted into secondary chemical-induced hyperglycaemic recipients. Maintenance of glucose tolerance required Rela but was independent of classical NF-κB inflammatory cascades, as blocking NF-κB signalling in vivo by beta cell knockout of Ikbkg (NEMO), or beta cell overexpression of Tnfaip3 (A20), did not cause severe glucose intolerance. Thus, basal p65 activity has an essential and islet-intrinsic role in maintaining normal glucose homeostasis. Genome-wide bioinformatic mapping revealed the presence of p65 binding sites in the promoter regions of specific metabolic genes and in the majority of islet enhancer hubs (~70% of ~1300 hubs), which are responsible for shaping beta cell type-specific gene expression programmes. Indeed, the islet-specific metabolic genes Slc2a2, Capn9 and Pfkm identified within the large network of islet enhancer hub genes showed dysregulated expression in βp65KO islets.Conclusions/interpretationThese data demonstrate an unappreciated role for RELA as a regulator of islet-specific transcriptional programmes necessary for the maintenance of healthy glucose metabolism. These findings have clinical implications for the use of anti-inflammatories, which influence NF-κB activation and are associated with diabetes.Graphical

Integrated Machine Learning and Bioinformatic Analyses Constructed a Network Between Mitochondrial Dysfunction and Immune Microenvironment of Periodontitis.

Periodontitis is a prevalent and persistent inflammatory condition that impacts the supporting tissues of the teeth, including the gums and bone. Recent research indicates that mitochondrial dysfunction may be involved in the onset and advancement of periodontitis. The current work sought to reveal the interaction between mitochondrial dysfunction and the immune microenvironment in periodontitis. Public data were acquired from MitoCarta 3.0, Mitomap, and GEO databases. Hub markers were screened out by five integrated machine learning algorithms and verified by laboratory experiments. Single-cell sequencing data were utilized to unravel cell-type specific expression levels of hub genes. An artificial neural network model was constructed to discriminate periodontitis from healthy controls. An unsupervised consensus clustering algorithm revealed mitochondrial dysfunction-related periodontitis subtypes. The immune and mitochondrial characteristics were calculated using CIBERSORTx and ssGSEA algorithms. Two hub mitochondria-related markers (CYP24A1 and HINT3) were identified. Single-cell sequencing data revealed that HINT3 was primarily expressed in dendritic cells, while CYP24A1 was mainly expressed in monocytes. The hub genes based artificial neural network model showed robust diagnostic performance. The unsupervised consensus clustering algorithm revealed two distinct mitochondrial phenotypes. The hub genes exhibited a strong correlation with the immune cell infiltration and mitochondrial respiratory chain complexes. The study identified two hub markers that may serve as potential targets for immunotherapy and provided a novel reference for future investigations into the function of mitochondria in periodontitis.

MDB-14. MULTI-OMICS OF HUMAN MEDULLOBLASTOMAS AND GENETIC LINEAGE TRACING IN VIVO IDENTIFY REGULATORS OF STEM-CELL MAINTENANCE

Abstract Recent studies implicate the glutamatergic lineage of the rhombic lip (RL) as the origin of group 3/4 medulloblastomas (MBs). MB stem cells resemble stalled RL progenitors and targeting common pluripotency factors induces differentiated phenotypes. However, the genetic and epigenetic regulators of stemness in MB remain incompletely understood. We profiled tissues resected from 38 patients who underwent surgery for MB at UCSF via single-nucleus RNA sequencing (snRNA-seq). Of these, 13 were also subjected to single-cell assay for transposase-accessible chromatin by sequencing (scATAC-seq). These assays generated 560,509 and 71,276 cells respectively, after quality control. Motif and latent-space analyses were used to infer cell-type specific transcription-factor expression and binding, and associated cis-regulatory enhancers. Further, we performed single-cell genetic lineage tracing on xenografts obtained from two distinct group-3/4 patient-derived lines, engineered to express heritable barcodes. We trained a machine-learning classifier on public snRNA-seq of human RL and classified Nestin-high quiescent stem-like cells, Nestin-moderate/ASPM-high proliferating progenitor cells, and glutamatergic lineage cells at various stages of differentiation in our MB clinical specimens and xenografts. Integrated analysis of these datasets maps the transition from quiescent to proliferating stem-like MB cells to more differentiated cell types at the level of transcription-factor activity and binding. We found that both the expression and binding of epigenetic regulators SUZ12, EZH2, DNMT1, and BRD7/8 were high in stem cells, but attenuated during lineage commitment. This was also the case for genes known to maintain neural stem cells in the developing brain, e.g. LIN28, FOXO3, HES6, LHX1/2/4, NUP98. Comparative analysis of developmental and MB gene expression, with motif analysis, implicates a conserved embryonal mechanism for indefinite replicative division in MB stem-like cells. These studies provide a single-cell atlas of transcriptomic and epigenetic cellular states in human MB, a map of MB cellular hierarchical organization and transcription-factor regulation of stem-cell maintenance in vivo.

Transcriptome-wide association study reveals novel susceptibility genes for coronary atherosclerosis.

Genetic risk factors substantially contributed to the development of coronary atherosclerosis. Genome-wide association study (GWAS) has identified many risk loci for coronary atherosclerosis, but the translation of these loci into therapeutic targets is limited for their location in non-coding regions. Here, we aimed to screen the potential coronary atherosclerosis pathogenic genes expressed though TWAS (transcriptome wide association study) and explore the underlying mechanism association. Four TWAS approaches (PrediXcan, JTI, UTMOST, and FUSION) were used to screen genes associated with coronary atherosclerosis. Enrichment analysis of TWAS-identified genes was applied through the Metascape website. The summary-data-based Mendelian randomization (SMR) analysis was conducted to provide the evidence of causal relationship between the candidate genes and coronary atherosclerosis. At last, the cell type-specific expression of the intersection genes was examined by using human coronary artery single-cell RNA-seq, interrogating the immune microenvironment of human coronary atherosclerotic plaque at different stages of maturity. We identified 19 genes by at least three approaches and 1 gene (NBEAL1) by four approaches. Enrichment analysis enriching the genes identified at least by two TWAS approaches, suggesting that these genes were markedly enriched in asthma and leukocyte mediated immunity reaction. Further, the summary-data-based Mendelian randomization (SMR) analysis provided the evidence of causal relationship between NBEAL1 gene and coronary atherosclerosis, confirming the protecting effects of NBEAL1 gene and coronary atherosclerosis. At last, the single cell cluster analysis demonstrated that NBEAL1 gene has differential expressions in macrophages, plasma cells and endothelial cells. Our study identified the novel genes associated with coronary atherosclerosis and suggested the potential biological function for these genes, providing insightful guidance for further biological investigation and therapeutic approaches development in atherosclerosis-related diseases.

Single-cell analysis of adult human heart across healthy and cardiovascular disease patients reveals the cellular landscape underlying SARS-CoV-2 invasion of myocardial tissue through ACE2

BackgroundThe distribution of ACE2 and accessory proteases (ANAD17 and CTSL) in cardiovascular tissue and the host cell receptor binding of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are crucial to understanding the virus’s cell invasion, which may play a significant role in determining the viral tropism and its clinical manifestations.MethodsWe conducted a comprehensive analysis of the cell type-specific expression of ACE2, ADAM17, and CTSL in myocardial tissue from 10 patients using RNA sequencing. Our study included a meta-analysis of 2 heart single-cell RNA-sequencing studies with a total of 90,024 cells from 250 heart samples of 10 individuals. We used co-expression analysis to locate specific cell types that SARS-CoV-2 may invade.ResultsOur results revealed cell-type specific associations between male gender and the expression levels of ACE2, ADAM17, and CTSL, including pericytes and fibroblasts. AGT, CALM3, PCSK5, NRP1, and LMAN were identified as potential accessory proteases that might facilitate viral invasion. Enrichment analysis highlighted the extracellular matrix interaction pathway, adherent plaque pathway, vascular smooth muscle contraction inflammatory response, and oxidative stress as potential immune pathways involved in viral infection, providing potential molecular targets for therapeutic intervention. We also found specific high expression of IFITM3 and AGT in pericytes and differences in the IFN-II signaling pathway and PAR signaling pathway in fibroblasts from different cardiovascular comorbidities.ConclusionsOur data indicated possible high-risk groups for COVID-19 and provided emerging avenues for future investigations of its pathogenesis.Trial registration(Not applicable).

Long Non-Coding RNAs, Cell Cycle, and Human Breast Cancer.

The long non-coding RNAs (lncRNAs) constitute an important class of the human transcriptome. The discovery of lncRNAs provided one of many unexpected results of the post-genomic era and uncovered a huge number of previously ignored transcriptional events. In recent years, lncRNAs are known to be linked with human diseases, with particular focus on cancer. Growing evidence has indicated that dysregulation of lncRNAs in breast cancer (BC) is strongly associated with the occurrence, development, and progress. Increasing numbers of lncRNAs have been found to interact with cell cycle progression and tumorigenesis in BC. The lncRNAs can exert their effect as a tumor suppressor or oncogene and regulate tumor development through direct or indirect regulation of cancer-related modulators and signaling pathways. What is more, lncRNAs are excellent candidates for promising therapeutic targets in BC due to the features of high tissue and cell-type specific expression. However, the underlying mechanisms of lncRNAs in BC still remain largely undefined. Here, we concisely summarize and sort out the current understanding of research progress in relationships of the roles for lncRNA in regulating the cell cycle. We also summarize the evidence for aberrant lncRNA expression in BC, and the potential for lncRNA to improve BC therapy is also discussed. Together, lncRNAs can be considered as exciting therapeutic candidates whose expression can be altered to impede BC progression.

Inference of cell type-specific gene regulatory networks on cell lineages from single cell omic datasets

Cell type-specific gene expression patterns are outputs of transcriptional gene regulatory networks (GRNs) that connect transcription factors and signaling proteins to target genes. Single-cell technologies such as single cell RNA-sequencing (scRNA-seq) and single cell Assay for Transposase-Accessible Chromatin using sequencing (scATAC-seq), can examine cell-type specific gene regulation at unprecedented detail. However, current approaches to infer cell type-specific GRNs are limited in their ability to integrate scRNA-seq and scATAC-seq measurements and to model network dynamics on a cell lineage. To address this challenge, we have developed single-cell Multi-Task Network Inference (scMTNI), a multi-task learning framework to infer the GRN for each cell type on a lineage from scRNA-seq and scATAC-seq data. Using simulated and real datasets, we show that scMTNI is a broadly applicable framework for linear and branching lineages that accurately infers GRN dynamics and identifies key regulators of fate transitions for diverse processes such as cellular reprogramming and differentiation.

Multimodal single cell analysis infers widespread enhancer co-activity in a lymphoblastoid cell line

Non-coding regulatory elements such as enhancers are key in controlling the cell-type specificity and spatio-temporal expression of genes. To drive stable and precise gene transcription robust to genetic variation and environmental stress, genes are often targeted by multiple enhancers with redundant action. However, it is unknown whether enhancers targeting the same gene display simultaneous activity or whether some enhancer combinations are more often co-active than others. Here, we take advantage of recent developments in single cell technology that permit assessing chromatin status (scATAC-seq) and gene expression (scRNA-seq) in the same single cells to correlate gene expression to the activity of multiple enhancers. Measuring activity patterns across 24,844 human lymphoblastoid single cells, we find that the majority of enhancers associated with the same gene display significant correlation in their chromatin profiles. For 6944 expressed genes associated with enhancers, we predict 89,885 significant enhancer-enhancer associations between nearby enhancers. We find that associated enhancers share similar transcription factor binding profiles and that gene essentiality is linked with higher enhancer co-activity. We provide a set of predicted enhancer-enhancer associations based on correlation derived from a single cell line, which can be further investigated for functional relevance.

A computational method for cell type-specific expression quantitative trait loci mapping using bulk RNA-seq data

Mapping cell type-specific gene expression quantitative trait loci (ct-eQTLs) is a powerful way to investigate the genetic basis of complex traits. A popular method for ct-eQTL mapping is to assess the interaction between the genotype of a genetic locus and the abundance of a specific cell type using a linear model. However, this approach requires transforming RNA-seq count data, which distorts the relation between gene expression and cell type proportions and results in reduced power and/or inflated type I error. To address this issue, we have developed a statistical method called CSeQTL that allows for ct-eQTL mapping using bulk RNA-seq count data while taking advantage of allele-specific expression. We validated the results of CSeQTL through simulations and real data analysis, comparing CSeQTL results to those obtained from purified bulk RNA-seq data or single cell RNA-seq data. Using our ct-eQTL findings, we were able to identify cell types relevant to 21 categories of human traits.

Associations of VGF with Neuropathologies and Cognitive Health in Older Adults.

VGF is proposed as a potential therapeutic target for Alzheimer's (AD) and other neurodegenerative conditions. The cell-type specific and, separately, peptide specific associations of VGF with pathologic and cognitive outcomes remain largely unknown. We leveraged gene expression and protein data from the human neocortex and investigated the VGF associations with common neuropathologies and late-life cognitive decline. Community-dwelling older adults were followed every year, died, and underwent brain autopsy. Cognitive decline was captured via annual cognitive testing. Common neurodegenerative and cerebrovascular conditions were assessed during neuropathologic evaluations. Bulk brain RNASeq and targeted proteomics analyses were conducted using frozen tissues from dorsolateral prefrontal cortex of 1,020 individuals. Cell-type specific gene expressions were quantified in a subsample (N = 424) following single nuclei RNASeq analysis from the same cortex. The bulk brain VGF gene expression was primarily associated with AD and Lewy bodies. The VGF gene association with cognitive decline was in part accounted for by neuropathologies. Similar associations were observed for the VGF protein. Cell-type specific analyses revealed that, while VGF was differentially expressed in most major cell types in the cortex, its association with neuropathologies and cognitive decline was restricted to the neuronal cells. Further, the peptide fragments across the VGF polypeptide resembled each other in relation to neuropathologies and cognitive decline. Multiple pathways link VGF to cognitive health in older age, including neurodegeneration. The VGF gene functions primarily in neuronal cells and its protein associations with pathologic and cognitive outcomes do not map to a specific peptide. ANN NEUROL 2023;94:232-244.