Abstract 17503: ETV2 Functions as a Pioneer Factor and Regulates the Endothelial Lineage

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Wuming Gong ◽  
Satyabrata Das ◽  
Javier Sierra-Pagan ◽  
Mary G Garry ◽  
Daniel J Garry
Keyword(s):  
Marker Genes ◽  
Cardiovascular Development ◽  
Rna Seq ◽  
Regulatory Pathways ◽  
Cardiovascular Regeneration ◽  
Multipotent Progenitors ◽  
Cellular Context ◽  
Pioneer Factor ◽  
Pioneer Factors ◽  
Endothelial Lineage

Background: Genetic mutations perturb the multipotent progenitors, which results in congenital cardiovascular disease. Therefore, it is essential to decipher the pioneer factors and the regulatory pathways that govern the specification and differentiation of mesodermal progenitors and use this information to develop targeted therapies to promote cardiovascular regeneration. Etv2 as an essential transcription factor for the development of cardiac, endothelial and hematopoietic lineages. In the present study, we used ES/EB differentiation and MEF reprogramming systems, to define Etv2 as a novel pioneer factor that relaxes the closed chromatin and drives endothelial development. Results: Using the iHA-Etv2 ES cell line, we engineered a mouse that inducibly overexpresses ETV2. The bulk RNA-seq, single cell RNA-seq data and ATAC-seq experiments showed that inducing Etv2 in MEFs and ES/EBs activated the downstream endothelial marker genes and promoted the development of endothelial lineages, supporting the notion that Etv2 functioned as a master regulator to drive the endothelial lineage development in different cellular contexts. We found that similar to other known pioneer factors, Etv2 was intrinsically able to target and bind the nucleosomes, and this capability appeared to be independent of the cellular context. To further define the mechanism, we performed Etv2, Brg1 and H3K27ac ChIP-seq analyses during MEF reprogramming and ES/EB differentiation. We found that Brg1 maintains and stabilizes the binding of Etv2 on the nucleosome, and Etv2 requires Brg1 to activate downstream genes during reprograming. Conclusion: In these studies, we defined Etv2 as a novel pioneer factor that relaxed the closed chromatin and promoted the endothelial lineage in both ES/EB differentiation and MEF reprogramming. The definition of these mechanisms will enhance our understanding of cardiovascular development and regeneration and serve as a platform for therapeutic applications for patients with congenital or aging related cardiovascular diseases.

scholarly journals Integrated genomic analysis reveals regulatory pathways and dynamic landscapes of the tRNA transcriptome

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Zefang Sun ◽  
Jia Tan ◽  
Minqiong Zhao ◽  
Qiyao Peng ◽  
Mingqing Zhou ◽  
...  
Keyword(s):  
Expression Profiles ◽  
Genomic Analysis ◽  
Rna Seq ◽  
Regulatory Pathways ◽  
Cellular Processes ◽  
Dynamic Landscapes ◽  
Rna Fragments ◽  
A Cell ◽  
Considerable Impact ◽  
New Algorithms

AbstracttRNAs and tRNA-derived RNA fragments (tRFs) play various roles in many cellular processes outside of protein synthesis. However, comprehensive investigations of tRNA/tRF regulation are rare. In this study, we used new algorithms to extensively analyze the publicly available data from 1332 ChIP-Seq and 42 small-RNA-Seq experiments in human cell lines and tissues to investigate the transcriptional and posttranscriptional regulatory mechanisms of tRNAs. We found that histone acetylation, cAMP, and pluripotency pathways play important roles in the regulation of the tRNA gene transcription in a cell-specific manner. Analysis of RNA-Seq data identified 950 high-confidence tRFs, and the results suggested that tRNA pools are dramatically distinct across the samples in terms of expression profiles and tRF composition. The mismatch analysis identified new potential modification sites and specific modification patterns in tRNA families. The results also show that RNA library preparation technologies have a considerable impact on tRNA profiling and need to be optimized in the future.

scholarly journals JIND: Joint Integration and Discrimination for Automated Single-Cell Annotation

2020 ◽  
Author(s):  
Mohit Goyal ◽  
Guillermo Serrano ◽  
Ilan Shomorony ◽  
Mikel Hernaez ◽  
Idoia Ochoa
Keyword(s):  
Single Cell ◽  
Cell Types ◽  
Marker Genes ◽  
Specific Marker ◽  
Rna Seq ◽  
Batch Effects ◽  
Cell Type ◽  
Latent Space ◽  
Cell Type Specific ◽  
Low Dimensional

AbstractSingle-cell RNA-seq is a powerful tool in the study of the cellular composition of different tissues and organisms. A key step in the analysis pipeline is the annotation of cell-types based on the expression of specific marker genes. Since manual annotation is labor-intensive and does not scale to large datasets, several methods for automated cell-type annotation have been proposed based on supervised learning. However, these methods generally require feature extraction and batch alignment prior to classification, and their performance may become unreliable in the presence of cell-types with very similar transcriptomic profiles, such as differentiating cells. We propose JIND, a framework for automated cell-type identification based on neural networks that directly learns a low-dimensional representation (latent code) in which cell-types can be reliably determined. To account for batch effects, JIND performs a novel asymmetric alignment in which the transcriptomic profile of unseen cells is mapped onto the previously learned latent space, hence avoiding the need of retraining the model whenever a new dataset becomes available. JIND also learns cell-type-specific confidence thresholds to identify and reject cells that cannot be reliably classified. We show on datasets with and without batch effects that JIND classifies cells more accurately than previously proposed methods while rejecting only a small proportion of cells. Moreover, JIND batch alignment is parallelizable, being more than five or six times faster than Seurat integration. Availability: https://github.com/mohit1997/JIND.

scholarly journals Comparative cellular analysis of motor cortex in human, marmoset and mouse

Nature ◽  
2021 ◽  
Vol 598 (7879) ◽  
pp. 111-119 ◽  
Author(s):  
Trygve E. Bakken ◽  
Nikolas L. Jorstad ◽  
Qiwen Hu ◽  
Blue B. Lake ◽  
Wei Tian ◽  
...  
Keyword(s):  
Motor Cortex ◽  
Primary Motor Cortex ◽  
Neuronal Cell ◽  
Cell Types ◽  
Morphological Characterization ◽  
Fine Motor ◽  
Marker Genes ◽  
Cell Type ◽  
Regulatory Pathways ◽  
Cellular Analysis

AbstractThe primary motor cortex (M1) is essential for voluntary fine-motor control and is functionally conserved across mammals1. Here, using high-throughput transcriptomic and epigenomic profiling of more than 450,000 single nuclei in humans, marmoset monkeys and mice, we demonstrate a broadly conserved cellular makeup of this region, with similarities that mirror evolutionary distance and are consistent between the transcriptome and epigenome. The core conserved molecular identities of neuronal and non-neuronal cell types allow us to generate a cross-species consensus classification of cell types, and to infer conserved properties of cell types across species. Despite the overall conservation, however, many species-dependent specializations are apparent, including differences in cell-type proportions, gene expression, DNA methylation and chromatin state. Few cell-type marker genes are conserved across species, revealing a short list of candidate genes and regulatory mechanisms that are responsible for conserved features of homologous cell types, such as the GABAergic chandelier cells. This consensus transcriptomic classification allows us to use patch–seq (a combination of whole-cell patch-clamp recordings, RNA sequencing and morphological characterization) to identify corticospinal Betz cells from layer 5 in non-human primates and humans, and to characterize their highly specialized physiology and anatomy. These findings highlight the robust molecular underpinnings of cell-type diversity in M1 across mammals, and point to the genes and regulatory pathways responsible for the functional identity of cell types and their species-specific adaptations.

scholarly journals Reprogramming of synovial macrophage metabolism by synovial fibroblasts under inflammatory conditions

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Noritaka Saeki ◽  
Yuuki Imai
Keyword(s):  
Gene Expression ◽  
Synovial Fibroblasts ◽  
Metabolic Reprogramming ◽  
Metabolic Status ◽  
Immunofluorescent Staining ◽  
Marker Genes ◽  
Rna Seq ◽  
Expression Levels ◽  
Inflammatory Conditions ◽  
Metabolic Genes

Abstract Background Macrophages adapt to microenvironments, and change metabolic status and functions to regulate inflammation and/or maintain homeostasis. In joint cavities, synovial macrophages (SM) and synovial fibroblasts (SF) maintain homeostasis. However, under inflammatory conditions such as rheumatoid arthritis (RA), crosstalk between SM and SF remains largely unclear. Methods Immunofluorescent staining was performed to identify localization of SM and SF in synovium of collagen antibody induced arthritis (CAIA) model mice and normal mice. Murine arthritis tissue-derived SM (ADSM), arthritis tissue-derived SF (ADSF) and normal tissue-derived SF (NDSF) were isolated and the purity of isolated cells was examined by RT-qPCR and flow cytometry analysis. RNA-seq was conducted to reveal gene expression profile in ADSM, NDSF and ADSF. Cellular metabolic status and expression levels of metabolic genes and inflammatory genes were analyzed in ADSM treated with ADSM-conditioned medium (ADSM-CM), NDSF-CM and ADSF-CM. Results SM and SF were dispersed in murine hyperplastic synovium. Isolations of ADSM, NDSF and ADSF to analyze the crosstalk were successful with high purity. From gene expression profiles by RNA-seq, we focused on secretory factors in ADSF-CM, which can affect metabolism and inflammatory activity of ADSM. ADSM exposed to ADSF-CM showed significantly upregulated glycolysis and mitochondrial respiration as well as glucose and glutamine uptake relative to ADSM exposed to ADSM-CM and NDSF-CM. Furthermore, mRNA expression levels of metabolic genes, such as Slc2a1, Slc1a5, CD36, Pfkfb1, Pfkfb3 and Irg1, were significantly upregulated in ADSM treated with ADSF-CM. Inflammation marker genes, including Nos2, Tnf, Il-1b and CD86, and the anti-inflammatory marker gene, Il-10, were also substantially upregulated by ADSF-CM. On the other hand, NDSF-CM did not affect metabolism and gene expression in ADSM. Conclusions These findings suggest that crosstalk between SM and SF under inflammatory conditions can induce metabolic reprogramming and extend SM viability that together can contribute to chronic inflammation in RA.

scholarly journals Single-nuclei RNA-seq on human retinal tissue provides improved transcriptome profiling

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Qingnan Liang ◽  
Rachayata Dharmat ◽  
Leah Owen ◽  
Akbar Shakoor ◽  
Yumei Li ◽  
...  
Keyword(s):  
Single Cell ◽  
Transcriptome Profiling ◽  
Cell Types ◽  
Retinal Cell ◽  
Peripheral Retina ◽  
Marker Genes ◽  
Rna Seq ◽  
Cell Type ◽  
Retinal Tissue ◽  
The Individual

AbstractSingle-cell RNA-seq is a powerful tool in decoding the heterogeneity in complex tissues by generating transcriptomic profiles of the individual cell. Here, we report a single-nuclei RNA-seq (snRNA-seq) transcriptomic study on human retinal tissue, which is composed of multiple cell types with distinct functions. Six samples from three healthy donors are profiled and high-quality RNA-seq data is obtained for 5873 single nuclei. All major retinal cell types are observed and marker genes for each cell type are identified. The gene expression of the macular and peripheral retina is compared to each other at cell-type level. Furthermore, our dataset shows an improved power for prioritizing genes associated with human retinal diseases compared to both mouse single-cell RNA-seq and human bulk RNA-seq results. In conclusion, we demonstrate that obtaining single cell transcriptomes from human frozen tissues can provide insight missed by either human bulk RNA-seq or animal models.

scholarly journals Transcriptomic Analysis Reveals Hormonal Control of Shoot Branching in Salix matsudana

Forests ◽  
2020 ◽  
Vol 11 (3) ◽  
pp. 287
Author(s):  
Juanjuan Liu ◽  
Bingbing Ni ◽  
Yanfei Zeng ◽  
Caiyun He ◽  
Jianguo Zhang
Keyword(s):  
Molecular Level ◽  
Hormonal Control ◽  
Molecular Techniques ◽  
Differentially Expressed ◽  
Shoot Branching ◽  
Rna Seq ◽  
Comprehensive Understanding ◽  
Regulatory Pathways ◽  
Salix Matsudana ◽  
Transcriptome Expression

Shoot branching is regulated by axillary bud activities, which subsequently grow into branches. Phytohormones play a central role in shoot branching control, particularly with regard to auxin, cytokinins (CKs), strigolactones (SLs), and gibberellins (GAs). To further study the molecular basis for the shoot branching in Salix matsudana, how shoot branching responds to hormones and regulatory pathways was investigated, and potential genes involved in the regulation of shoot branching were identified. However, how these positive and inhibitory processes work on the molecular level remains unknown. RNA-Seq transcriptome expression analysis was used to elucidate the mechanisms underlying shoot branching. In total, 102 genes related to auxin, CKs, SLs, and GAs were differentially expressed in willow development. A majority of the potential genes associated with branching were differentially expressed at the time of shoot branching in S. matsudana, which have more number of branching. These findings are consistent with the growth and physiological results. A regulatory network model was proposed to explain the interaction between the four hormones that control shoot branching. Collectively, the results presented here contribute to a more comprehensive understanding of the hormonal effects on shoot branching in S. matsudana. In the future, these findings will help uncover the interactions among auxin, SLs, CKs, and GAs that control shoot branching in willow, which could help improve plant structures through the implementation of molecular techniques in targeted breeding.

scholarly journals PPARG Could Work as a Valid Therapeutic Strategy for the Treatment of Lung Squamous Cell Carcinoma

PPAR Research ◽  
2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Shunbin Shi ◽  
Guiping Yu ◽  
Bin Huang ◽  
Yedong Mi ◽  
Yan Kang ◽  
...  
Keyword(s):  
Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Squamous Cell ◽  
Lung Squamous Cell Carcinoma ◽  
Nonsmall Cell Lung Cancer ◽  
Ppar Gamma ◽  
Gene Set Enrichment Analysis ◽  
Marker Genes ◽  
Molecular Pathways ◽  
Regulatory Pathways

Previous studies showed that PPAR-gamma (PPARG) ligands might serve as potential therapeutic agents for nonsmall cell lung cancer (NSCLC). However, a few studies reported the specific relationship between PPARG and lung squamous cell carcinoma (LSCC). Here, we made an effort to explore the relationship between PPARG and LSCC. First, we used mega-analysis and partial mega-analysis to analyze the effects of PPARG on LSCC by using 12 independent LSCC expression datasets (285 healthy controls and 375 LSCC cases). Then, literature-based molecular pathways between PPARG and LSCC were established. After that, a gene set enrichment analysis (GSEA) was conducted to study the functionalities of PPARG and PPARG-driven triggers within the molecular pathways. Finally, another mega-analysis was constructed to test the expression changes of PPARG and its driven targets. The partial mega-analysis showed a significant downregulated expression of PPARG in LSCC (LFC=−1.08, p value=0.00073). Twelve diagnostic markers and four prognostic markers were identified within multiple PPARG-LSCC regulatory pathways. Our results suggested that the activation of PPARG expression may inhibit the development and progression of LSCC through the regulation of LSCC upstream regulators and downstream marker genes, which were involved in tumor cell proliferation and protein polyubiquitination/ubiquitination.

scholarly journals MiMIF-2 Effector of Meloidogyne incognita Exhibited Enzyme Activities and Potential Roles in Plant Salicylic Acid Synthesis

2020 ◽  
Vol 21 (10) ◽  
pp. 3507
Author(s):  
Jianlong Zhao ◽  
Zhenchuan Mao ◽  
Qinghua Sun ◽  
Qian Liu ◽  
Heng Jian ◽  
...  
Keyword(s):  
Salicylic Acid ◽  
Transcriptome Analysis ◽  
Meloidogyne Incognita ◽  
Plant Immunity ◽  
Acid Synthesis ◽  
Host Immunity ◽  
Parasitic Nematodes ◽  
Marker Genes ◽  
Rna Seq ◽  
Shock Proteins

Plant-parasitic nematodes secrete a series of effectors to promote parasitism by modulating host immunity, but the detailed molecular mechanism is ambiguous. Animal parasites secrete macrophage migration inhibitory factor (MIF)-like proteins for evasion of host immune systems, in which their biochemical activities play essential roles. Previous research demonstrated that MiMIF-2 effector was secreted by Meloidogyne incognita and modulated host immunity by interacting with annexins. In this study, we show that MiMIF-2 had tautomerase activity and protected nematodes against H2O2 damage. MiMIF-2 expression not only decreased the amount of H2O2 generation during nematode infection in Arabidopsis, but also suppressed Bax-induced cell death by inhibiting reactive oxygen species burst in Nicotiana benthamiana. Further, RNA-seq transcriptome analysis and RT-qPCR showed that the expression of some heat-shock proteins was down regulated in MiMIF-2 transgenic Arabidopsis. After treatment with flg22, RNA-seq transcriptome analysis indicated that the differentially expressed genes in MiMIF-2 expressing Arabidopsis were pointed to plant hormone signal transduction, compound metabolism and plant defense. RT-qPCR and metabolomic results confirmed that salicylic acid (SA) related marker genes and SA content were significantly decreased. Our results provide a comprehensive understanding of how MiMIF-2 modulates plant immunity and broaden knowledge of the intricate relationship between M. incognita and host plants.

scholarly journals GAF is essential for zygotic genome activation and chromatin accessibility in the early Drosophila embryo

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Marissa M Gaskill ◽  
Tyler J Gibson ◽  
Elizabeth D Larson ◽  
Melissa M Harrison
Keyword(s):  
Chromatin Accessibility ◽  
Drosophila Embryo ◽  
Gaga Factor ◽  
Zygotic Transcription ◽  
Early Drosophila Embryo ◽  
Zygotic Gene ◽  
Pioneer Factor ◽  
Pioneer Factors ◽  
Genome Activation ◽  
Development Proceeds

Following fertilization, the genomes of the germ cells are reprogrammed to form the totipotent embryo. Pioneer transcription factors are essential for remodeling the chromatin and driving the initial wave of zygotic gene expression. In Drosophila melanogaster, the pioneer factor Zelda is essential for development through this dramatic period of reprogramming, known as the maternal-to-zygotic transition (MZT). However, it was unknown whether additional pioneer factors were required for this transition. We identified an additional maternally encoded factor required for development through the MZT, GAGA Factor (GAF). GAF is necessary to activate widespread zygotic transcription and to remodel the chromatin accessibility landscape. We demonstrated that Zelda preferentially controls expression of the earliest transcribed genes, while genes expressed during widespread activation are predominantly dependent on GAF. Thus, progression through the MZT requires coordination of multiple pioneer-like factors, and we propose that as development proceeds control is gradually transferred from Zelda to GAF.

scholarly journals Folic Acid Exerts Dose-Dependent Biphasic Effects on Cardiac Development of Zebrafish Embryos

2021 ◽  
Author(s):  
Xuhui Han ◽  
Bingqi Wang ◽  
Hongjie Wang ◽  
Yao Zu
Keyword(s):  
Folic Acid ◽  
Methylenetetrahydrofolate Reductase ◽  
Cardiac Development ◽  
Mthfr Gene ◽  
Folate Metabolism ◽  
Marker Genes ◽  
Zebrafish Embryos ◽  
Cardiovascular Development ◽  
Knock Out ◽  
Dose Dependent

Folic acid, one of the 13 essential vitamins, plays an important role in cardiovascular development. Mutations in folic acid synthesis gene 5,10-methylenetetrahydrofolate reductase (MTHFR) is significantly associated with the occurrence of congenital heart disease. However, the mechanisms underlying the regulation of cardiac development by mthfr gene are poorly understood. Here, we exposed zebrafish embryos to excessive folate or folate metabolism inhibitors. And we established a knock-out mutant of mthfr gene in zebrafish by using CRISPR/Cas9. The zebrafish embryos of insufficient or excessive folic acid, and mthfr-/- mutant all gave rise to early pericardial edema and cardiac defect at 3 days after fertilization(dpf). Furthermore, the folic acid treated embryos showed abnormal movement at 5dpf. The expression levels of cardiac marker genes hand2, gata4 and nppa changed in the abnormality of folate metabolism embryos and mthfr-/- mutant, and there is evidence that they are related to the change of methylation level caused by the change of folate metabolism. In conclusion, our study provides a novel model for the in-depth study of MTHFR gene and folate metabolism. And our results reveal that folic acid has a dose-dependent biphasic effect on early cardiac development.

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