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: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: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: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:A subset of transcription factors is proposed to form functional interactions with RNA to facilitate proper regulation of gene expression. Sox2, a critical transcription factor for maintenance of pluripotency and neurogenesis, is found associated with several lncRNAs, although it is unknown whether these interactions are direct or via other proteins. We demonstrate that human Sox2 interacts directly with one of these lncRNAs with high affinity through its HMG DNA-binding domain. Furthermore, rather than interacting with a single high affinity site, Sox2 binds regions of dsRNA in a non-sequence specific fashion. RNA binding is competitive with dsDNA, yet interaction with RNA is primarily electrostatically driven using a partially overlapping set of amino acids as that used for DNA binding. Finally, we use two distinct crosslinking techniques, formaldehyde and UV, with RNA immunoprecipitation (respectively, “fRIP” and “UVRIP”) of Sox2 in mouse embryonic stem cells to identify >1000 potential Sox2-RNA interacting partners with ~75% overlap between the two techniques. Together, these data reveal that Sox2 productively binds dsRNA with affinities rivaling ChIP-validated promoter DNA sites, suggesting that the transcriptome plays a significant role in regulation of gene expression by Sox2.
