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Single-Cell Sequencing Reveals Lineage-Specific Dynamic Genetic Regulation of Gene Expression During Human Cardiomyocyte Differentiation

ABSTRACT: Single-Cell Sequencing Reveals Lineage-Specific Dynamic Genetic Regulation of Gene Expression During Human Cardiomyocyte Differentiation

PROVIDER: PRJNA733138 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14667135_1.fastq.gz	Fastqsanger.gz
	SRR14667135_2.fastq.gz	Fastqsanger.gz
	SRR14667136_1.fastq.gz	Fastqsanger.gz
	SRR14667136_2.fastq.gz	Fastqsanger.gz
	SRR14667137_1.fastq.gz	Fastqsanger.gz

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Single-cell sequencing reveals lineage-specific dynamic genetic regulation of gene expression during human cardiomyocyte differentiation.

Project description:Dynamic and temporally specific gene regulatory changes may underlie unexplained genetic associations with complex disease. During a dynamic process such as cellular differentiation, the overall cell type composition of a tissue (or an in vitro culture) and the gene regulatory profile of each cell can both experience significant changes over time. To identify these dynamic effects in high resolution, we collected single-cell RNA-sequencing data over a differentiation time course from induced pluripotent stem cells to cardiomyocytes, sampled at 7 unique time points in 19 human cell lines. We employed a flexible approach to map dynamic eQTLs whose effects vary significantly over the course of bifurcating differentiation trajectories, including many whose effects are specific to one of these two lineages. Our study design allowed us to distinguish true dynamic eQTLs affecting a specific cell lineage from expression changes driven by potentially non-genetic differences between cell lines such as cell composition. Additionally, we used the cell type profiles learned from single-cell data to deconvolve and re-analyze data from matched bulk RNA-seq samples. Using this approach, we were able to identify a large number of novel dynamic eQTLs in single cell data while also attributing dynamic effects in bulk to a particular lineage. Overall, we found that using single cell data to uncover dynamic eQTLs can provide new insight into the gene regulatory changes that occur among heterogeneous cell types during cardiomyocyte differentiation.

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Single-Cell Sequencing Reveals Lineage-Specific Dynamic Genetic Regulation of Gene Expression During Human Cardiomyocyte Differentiation

Project description:The impact of genetic regulatory elements on gene expression can vary across cell states. During dynamic processes such as cellular differentiation, cells transition through multiple cell states, and may differentiate toward multiple terminal cell types. We collected time-series single-cell RNA-sequencing data from 19 human cell lines, capturing 7 time points along a 16-day differentiation from induced pluripotent stem cells to cardiomyocytes. We used unsupervised clustering, marker gene expression patterns, and pseudotime inference methods to map individual cells to a position along one of two bifurcating differentiation trajectories that were inferred from the data. We then identified genetic effects on gene regulation with varying effects across these trajectories. We identified hundreds of dynamic eQTLs that change significantly across pseudotime, including many variants whose effects are specific to one of the two lineages. We then re-analyzed previously collected bulk data, and used cell state information to infer cell type interaction eQTLs in bulk, assigning previously identified dynamic eQTLs to one of the newly characterized cellular trajectories.

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Dynamic cohesin is required for lineage specific distal gene regulation

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Project description:The cohesin complex plays essential roles in sister chromatin cohesin, chromosome organization and gene expression. The role it plays in the regulation of genes is incompletely understood. Here, we report that the cohesin release factor WAPL in mouse embryonic stem cells is crucial for maintaining a pool of dynamically bound cohesin to regions in the genome that are associated with lineage specific genes. These regions are enriched for active enhancer marks and transcription factor binding sites. Stabilization of cohesin, which leads to a loss of dynamic cohesin from these regions does not affect transcription factor binding or active enhancer marks, but does result in changes in promoter-enhancer interactions and downregulation of genes. Acute cohesin depletion can phenocopy the effect of WAPL depletion, showing that cohesin plays a crucial role in maintaining expression of lineage specific genes. The binding of dynamic cohesin to chromatin is dependent on the pluripotency transcription factor OCT4, but not NANOG. Finally, sites of dynamic cohesin binding sites are also found in differentiated cells, suggesting that they represent a general regulatory principle. We propose that the binding of dynamic cohesin to regulatory sites creates a favorable spatial environment in which promoters and enhancers can communicate to ensure proper gene expression.

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Project description:The differentiation to cardiomyocytes is a prerequisite and an important part of heart development. A good understanding of the complicated cardiomyocyte differentiation process benefits cardiogenesis study. Embryonic stem cells (ESCs), cell lines with infinite ability to proliferate and to be differentiated into all cell types of the adult body, are important research tools for investigation of differentiation and meanwhile good models for developmental research. In the current study, genome-wide gene expression of ESCs is profiled through high throughput platform during cardiomyocyte-specific differentiation and maturation. Gene expression patterns of undifferentiated ESCs and ESC-derived cardiomyocytes provide a global overview of genes involved in cardiomyocyte-specific differentiation, whereas marker gene expression profiles of both ESC-related genes and cardiac-specific genes presented the expression pattern shift during differentiation in a pure ESC-derived cardiomyocyte cell culture system. Transgenic mouse ESC clone with M-NM-1-MHCM-bM-^@M-^SPacM-bM-^@M-^SIRESM-bM-^@M-^SEGFP vector containing the EGFP gene and the PuromycinR (Pac) cassette under control of the cardiac M-NM-1-myosin heavy chain (M-NM-1-MHC) promoter was cultured and induced into differentiation. Puromycin was applied after differentiation start to enable cardiomyocyte-specific differentiation. Cells were harvested at 4 time points after differentiation start (day0, day12, day19 and day26). Two biological replicates were taken for every time point.

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CHD-associated enhancers direct human cardiomyocyte lineage commitment.

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