GENOMICS- Dissecting Temporal Enhancer Dynamics in Mouse Limb Development

This study explores the molecular mechanisms controlling transcriptional onset and maintenance of developmental genes during organogenesis. Utilizing the mouse limb as a model system, we aim to produce a high-resolution map of early-acting enhancers, termed initiating elements, and late-acting enhancers, termed maintenance elements. To study initiating elements, we will use a Sall4-marking approach to tag limb progenitors and their daughter cells. We will sort these cells and generate H3K27ac CUT&RUN maps to identify regulatory regions. To study maintenance elements, we will combine cell-type sorting in late-developing E14.5 limbs with H3K27ac CUT&RUN, along with 10x Multiome profiling of Sall4-descendant cells in late E14.5 limbs. By integrating these late-stage datasets, we will generate a comprehensive and cell-type-specific map of maintenance element. By combining the initiating and maintenance element maps over time, we will identify the temporal dynamics of enhancers regulating limb patterning genes. Specific enhancer combinations will then be deleted to investigate the roles of initiating and maintenance elements in shaping expression patterns. This project will provide the first temporally resolved and cell-type-specific atlas of limb enhancers, offering novel insights into how enhancers establish and maintain gene expression patterns over time.