UC Davis

The genomic sequence of the horse has been available since 2007, providing critical resources for discovering important genomic variants regarding both animal health and population structure. However, to fully understand the functional implications of these variants, detailed annotation of the horse genome is required. Currently, the horse genome is annotated using the limited available RNA-seq data, as well as through comparative genomics by translating human and mouse genome annotation. While this approach has served the equine researchers well and led to a number of discoveries improving the care and management of horses, many important questions remain unanswered. The limitation of the current annotation is two pronged. First, a comparative genomics approach is insufficient to identify many genes that are less evolutionarily conserved, especially those that are noncoding. The sole reliance on short-read RNA-seq data also meant that alternate isoforms could not be accurately resolved. Second, epigenomic regulatory elements are crucial to detailed understanding of gene expression network but are yet to be systemically identified in the horse. Many regulatory elements, including enhancers, promoters, and insulators, are not transcribed or transcribed at a very low level, necessitating alternate approaches to identify them. To solve these problems, the Functional Annotation of the Animal Genomes (FAANG) project proposed a systemic approach to tissue collection, phenotyping, and data generation, adopting the blueprint laid out by the Encyclopedia of DNA Elements (ENCODE) project. This thesis describes the equine FAANG team’s effort to map tissue-specific gene expression and regulation in the horse genome. Chapter 1 provides an overview of the equine FAANG project’s approach to functional annotation. Chapter 2 describes an improved transcriptome that includes novel genes and alternate isoforms compared to the current annotation. In Chapter 3, we use ATAC-seq to create a catalog of tissue-specific open chromatin regions, which can serve as proxies to active regulatory elements. Chapter 4 provides a complete annotation of chromatin states across nine tissues. The Addendum detailed our effort to validate assays for transposase accessible chromatin using sequencing (ATAC-seq) in both frozen tissues and cryopreserved nuclei from fresh tissues. This thesis presents the first comprehensive overview of gene expression and their regulation in the horse, enabling interrogation of complex gene regulatory network and further studies of complex traits in horses. Future work should focus on both widening the scope of the equine FAANG project by including more tissue types and developmental stages, as well as refining gene network at single-cell resolution.