Adult teleost fish and urodele amphibians can regenerate entire amputated appendages. By contrast, regenerative healing of adult mammalian limbs is limited to the very tips of digits. One of the key challenges in developmental biology is to understand how and why tissue regeneration occurs. The hallmark of limb or fin regeneration is formation of a blastema, a mesenchymal structure that contains progenitor cells for new skeletal elements. As regeneration proceeds, blastemal cell proliferation and patterning are regulated such that lost tissues of correct size and shape are replaced. While the catalogue of defined cell dynamics and molecular factors in tissue regeneration is expanding, we know much less of how genes involved in regenerative events are engaged upon injury. That is, what are the DNA sequences that recruit activators (or repressors) of gene programs during tissue regeneration, how are these sequences distributed throughout the genomes of regeneration-competent animals, and what are the transcription factors that engage with these sequences? Recently, we identified a class of distal gene regulatory elements that preferentially activate gene expression during regeneration, can be engineered in simple constructs to express developmental factors that promote regeneration, and have the potential to be recognized by the transcriptional machinery of distant species. The overall goal of this proposal is to discover gene regulatory concepts and mechanisms that restore size and pattern to an amputated appendage. We will test the hypothesis that enhancer and silencer elements are crucial regulatory modules enabling appendage regeneration in zebrafish. This work will increase understanding of developmental regulation during vertebrate tissue regeneration, and provide important perspective for comprehending, and perhaps changing, the existing limitations in regenerative capacity of human tissues.