Organismal physiology is built upon the foundation of molecular processes. A central axis to maintaining homeostasis in vivo is at the level of gene regulation. Tissue specific gene expression is created at the level of epigenetics, where proteins guided by tissue specific DNA binding proteins create a chromatin landscape for precise gene programs. Understanding these molecular processes is of vital importance to understand the underpinning pathologies, such as metabolic syndrome, which are a growing medical concern and require greater research efforts in order to tackle its challenges. A major epigenetic regulator is histone deacetylase 3 (HDAC3), which is a core member of the nuclear receptor corepressor (NCoR) complex. This ubiquitously expressed chromatin associated protein complex functions to repress target gene transcription. Here we address the functional role of HDAC3 in β-cells of adult mice. An HDAC3 β-cell specific knockout was generated using the MIP-CreERT transgenic mouse model and while HDAC3 β-cell specific deletion did not increase total pancreatic insulin content, the mice demonstrated markedly improved glucose tolerance and increased glucose-stimulated insulin secretion. Cistromic and transcriptomic analyses of pancreatic islets revealed that HDAC3 regulated multiple genes that contribute to glucose-stimulated insulin secretion. Furthermore, using mass spectrometry in conjunction of cistromic analyses of interactors we have characterized the interactome of HDAC3 and detailed its function in mammalian liver. We identified PROX1 as an abundant interactor which is corecruited with HDAC3 by HNF4a in liver to corepress gene transcription important for maintenance of lipid homeostasis. Lastly, as we continue to explore the protein-protein interaction networks of these critical factors, novel tools are proving to be invaluable to their investigation. The advent of CRISPR-Cas9 genome editing has allowed for reliable and simple design and generation of mouse models. Therefore we have employed this technology to generate a variety of epitope tagged mouse models with the goal of comparing their tissue specific interactomes. This body of work includes a wide breadth of biological techniques that have succeeded in advancing knowledge of HDAC3 function in vivo, vital to our understanding of molecular pathology in diabetes and obesity.