Browsing by Subject "hESCs"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Dynamic Changes in DNA Replication Timing and 3D Genome Organization During Cardiac Differentiation
    (2023-03) Martinez Cifuentes, Santiago
    Genome architecture has emerged as a key factor of gene regulation and is tightly coordinated with the temporal order of DNA replication (Replication Timing – RT). RT and 3D genome organization change dynamically throughout development in correlation with the establishment of cell type specific gene expression patterns. The first organ to develop during embryonic development is the heart. The heart is a post-mitotic, terminally differentiated organ which, compared to other organs such as the liver, lacks the capacity to regenerate upon injury such as myocardial infarction. Instead, it forms fibrotic tissue that maintains organ integrity but undermines pump function, often leading to congestive heart failure and premature death. Deciphering the 3D genome organization in cardiac differentiation may help our understanding of gene regulation during cardiac development. DNA replication timing (RT) is a very informative functional readout of large-scale chromatin organization across distinct cell types and its regulation during development. RT is cell type-specific, highly conserved and changes in RT affect approximately half the genome during development and differentiation. I hypothesize that changes in the 3D genome organization play a critical role in gene regulation and cell function in cardiac differentiation. In this work, I will differentiate human embryonic stem cells (hESCs) towards cardiomyocytes and use a multiomics approach (RT, transcriptome and 3D genome organization) to identify the dynamic changes in genome architecture, RT and gene expression during normal cardiac development. This will allow us, in the future, to construct an integrative model of nuclear function in cardiac cells that can be leveraged as a framework to identify cellular alterations associated with congenital cardiovascular diseases.
  • Thumbnail Image
    Item
    Dynamic regulation of replication timing during cell lineage decisions
    (2023-12) Sadu Murari, Lakshana Sruthi
    During embryogenesis, the fetal liver is one of the first organs to develop after the heart, relying heavily on signals from the adjacent cardiac mesoderm. As lineage specification proceeds during development and cells differentiate, genomic regions are segregated into early or late replicating domains, establishing a cell type-specific replication timing (RT) program. Decades of intensive research have uncovered a strong correlation between developmental changes in RT, genome organization, and the transcriptome. While global changes in RT have been identified during distinct differentiation pathways, lineage-specific changes during organ development remain unexplored. Identifying factors regulating changes in replication timing pertaining to cardiac and hepatic development would help better understand how replication timing is regulated during lineage decisions and help establish better models to study congenital defects. Here, we sought to study the developmental switches in RT, and the corresponding changes in the transcriptome, of hESC-derived hepatocyte-like cells and cardiomyocyte-like cells. We also developed a novel multi-omics approach for the parallel analysis of replication timing and gene expression (PARTAGE), as well as an optimized method for live single-cell subcloning of HepG2 cells to enable genome editing experiments and generation of mutant cell lines.