Project description:RNA-seq data indicated that Knockdown of GATA4 in human cardiomyocytes resulted in differential alternative splicing changes in genes involved in cytoskeleton organization and calcium ion import. Enhanced crosslinking and immunoprecipitation (CLIP) assay demonstrated that GATA4 directly binds to defined mRNAs motifs in a sequence-specific manner
Project description:Heterozygous mutations in GATA4 cause congenital heart defects and cardiomyopathy through unknown mechanisms. To gain insights into the trancriptome perturbations during human cardiac development due to GATA4 heterozygosity, we performed RNA-seq of isogenic wildtype and GATA4-G296S diseased cardiac progenitors (CPCs) and cardiomyocytes (CMs).
Project description:Using a systems biology approach, we discovered and dissected a three-way interaction between the immune system, the intestinal epithelium, and the microbiota. We found that mice lacking B lymphocytes, or lacking IgA, have low intestinal expression of lipid metabolism genes regulated by the transcription factor GATA4, and a consequent decrease in fat absorption in the intestine. The defect disappeared in germ free mice, suggesting that it is dependent on the microbiota; and sequencing analysis of the bacteria showed subtle differences between normal and B-cell deficient mice. Analysis of gene expression of gut biopsies from patients with common variable immunodeficiency and intestinal dysfunction revealed a high similarity to mouse B-cell knockout profiles. These data provide an explanation for a longstanding enigmatic association between immunodeficiency and defective lipid absorption in humans. This series represents the subsection of the study where we address the role of transcription factor Gata4. The data are from conditional KO (Gata4KOvil) and corresponding control mice.
Project description:Certain transcription factors are proposed to form functional interactions with RNA to facilitate proper regulation of gene expression. Sox2, a transcription factor critical for maintenance of pluripotency and neurogenesis, has been found associated with several lncRNAs, although it is unknown whether these interactions are direct or via other proteins. Here we demonstrate that human Sox2 interacts directly with one of these lncRNAs with high affinity through its HMG DNA-binding domain in vitro. These interactions are primarily with double-stranded RNA in a non-sequence specific fashion, mediated by a similar but not identical interaction surface. We further determined that Sox2 directly binds RNA in mouse embryonic stem cells by UV-cross-linked immunoprecipitation of Sox2 and more than a thousand Sox2-RNA interactions in vivo were identified using fRIP-seq. Together, these data reveal that Sox2 employs a high-affinity/low-specificity paradigm for RNA binding in vitro and in vivo.
Project description:Heterozygous mutations in GATA4 cause congenital heart defects and cardiomyopathy through unknown mechanisms. To gain insights into the open chromatin status during human cardiac development due to GATA4 heterozygosity, we performed ATAC-seq of wildtype and GATA4-G296S diseased cardiac progenitors.
Project description:Heterozygous mutations in GATA4 cause congenital heart defects and cardiomyopathy through unknown mechanisms. To gain insights into the genome-wide localization perturbations during human cardiac development due to GATA4 heterozygosity, we performed ChIP-seq of wildtype and GATA4-G296S diseased cardiomyocytes.
Project description:DNA methylation changes dynamically during development and is essential for embryogenesis in mammals. However, how DNA methylation affects developmental gene expression and cell differentiation remains elusive. During embryogenesis, many key transcription factors are used repeatedly, triggering different outcomes depending on the cell type and developmental stage. Here, we report that DNA methylation modulates transcription-factor output in the context of cell differentiation. Using a drug-inducible Gata4 system and a mouse embryonic stem (ES) cell model of mesoderm differentiation, we examined the cellular response to Gata4 in ES and mesoderm cells. The activation of Gata4 in ES cells is known to drive their differentiation to endoderm. We show that the differentiation of wild-type ES cells into mesoderm blocks their Gata4-induced endoderm differentiation, while mesoderm cells derived from ES cells that are deficient in the DNA methyltransferases Dnmt3a and Dnmt3b can retain their response to Gata4, allowing lineage conversion from mesoderm cells to endoderm in part. Transcriptome analysis of the cells' response to Gata4 over time revealed groups of endoderm and mesoderm developmental genes whose expression was induced by Gata4 only when DNA methylation was lost, suggesting that DNA methylation restricts the ability of these genes to respond to Gata4, rather than controlling their transcription per se. Our data indicate that epigenetic regulation by DNA methylation functions as a heritable safeguard to prevent transcription factors from activating inappropriate downstream genes, thereby contributing to the restriction of the differentiation potential of somatic cells.
