Defining the mechanism of chromatin accessibility modifications in vertebrate appendage regeneration

University Of Washington, Seattle WA

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Abstract

PROJECT SUMMARY Amphibians are able to undergo scarless healing and regeneration in response to amputation of their appendages. By articulating the cell and molecular basis for this regenerative capacity, we can develop therapeutic strategies to improve regenerative healing in humans. Successful regeneration in amphibians requires the early action of histone deacetylases (HDACs), enzymes that act to condense chromatin and inhibit transcription. However, the genomic and transcriptional targets of HDAC activity are unknown in this context, limiting our ability to form a mechanistic model of epigenetic and transcriptional reprogramming in vertebrate regeneration. It has been difficult to identify these targets because there are few resources for querying chromatin structure in regenerating vertebrate tissue. We have overcome this barrier by using a new assay for transposase accessible chromatin (ATAC-Seq) in Xenopus tropicalis tail regeneration. We have found that thousands of promoter regions are rapidly rendered inaccessible at 6 hours post amputation: the very same time that HDAC activity is required. These regions are later re-opened. Therefore, the central hypothesis of our study is that HDAC activity acts transiently to condense chromatin in these promoter regions by deacetylating specific core histone residues, and is reversed by the later action of histone acetyltransferases (HATs). Here will use our expertise in genomics and functional perturbations of this system to conduct a succinct functional secondary analysis that will test this hypothesis. We will identify the genomic regions that are sensitive to HDAC and HAT activity, the specific residues that are targeted, and the spatiotemporal distribution of histone acetylation in the regenerating tail. Importantly, we will also lay the foundation for future work that will establish how HDAC activity is balanced with other chromatin remodeling activities to reprogram transcription and cell behavior early in regeneration. !