Cellular states are governed by complex interactions involving transcription factors and DNA regulatory elements that drive expression of genes. At the single-cell level, DNA regulatory elements exist in two copies and are subjected to considerable stochasticity in their regulation. Stochastic epigenetic processes are fundamental to our understanding of gene regulation; however, little is known regarding i) how epigenetic states in single-cells vary through time, ii) how chromatin factors may promote variability of regulatory elements, iii) how these noisy regulatory elements may lead to functional differences in expression and iv) how this variance affects cellular decisions. Only recently have methods to measure chromatin states in single-cells become possible, providing a unique opportunity to measure epigenomic dynamics enabling mechanistic studies of epigenomic variability and their function in cells. Here, we detail these advances and propose to use these tools to study mechanisms of epigenetic memory, here defined as the inheritance of chromatin states across cell division. As the goal of this work, we seek to understand how these epigenomic alterations may alter lineage potential in hematopoietic stem cells in normal and diseased states, and also propose to expand the single-cell epigenomic toolbox for exploring new aspects of gene regulation. Throughout these efforts we will generate rich data sets and computational tools to provide a generalizable platform for unraveling cis- and trans-effectors of cellular differentiation during health and disease. Altogether, this work will provide new insight into gene regulation at the single-cell level and provide a powerful and broadly applicable toolbox for regulatory investigations of single-cells.
Traditional methods used to study the epigenome, the structure and modifications of DNA that regulate our genes, lack the resolution to measure epigenomic dynamics in human tissues. This proposal aims to use hematopoiesis as a model for understanding how persistent changes in epigenetic states, or `epi-mutations' can affect gene expression and cell fate decisions in healthy and diseased cells. We also describe our efforts to expand our experimental toolkit, which will empower research efforts that seek to understand regulatory changes across diverse cell types, which may ultimately help guide the direction of therapeutics that seek to intervene in disease progression.
