Date of Award
Doctor of Philosophy (PhD)
Cohen Paul Laboratory
Adipose tissue is a complex organ that is essential for lipid storage and energy homeostasis in mammals. While transcription factors that govern adipogenesis, inflammation, non-shivering thermogenesis, and endocrine function in fat tissue have been well-described, comparatively less attention has been given to post-transcriptional regulators. Here, we employ two contrasting approaches to identify miRNAs that influence adipocyte phenotype. First, miRNAs were sequenced from 6 fat pads, primary preadipocytes, and primary mature adipocytes. Comparative analyses of abundance across adipose tissue type and stage of differentiation revealed a number of miRNAs that influence key metabolic processes. miR-335 inhibited adipogenesis, miR-192 promoted pro-thermogenic and pro-inflammatory pathways, and miR-338 upregulated a variety of genes involved in both lipogenesis and lipolysis/β-oxidation. Although most effects on mRNA abundance were modest, these candidates and others identified in the analysis provide a foundation for further study. The second portion of this thesis focuses on network-based approaches to describe miRNA regulators in adipose tissue with an emphasis on miRNA binding activity. Most notably, we used Ago HITS-CLIP to comprehensively map mRNA:miRNA interactions in brown and white fat, revealing 21,281 unique miRNA binding sites in 6,717 genes. Most binding sites were shared between brown and white fat, although reads per binding site and reads per gene varied substantially. Targets for each miRNA were ranked to generate a catalog of miRNA binding activity. The miR-29 family emerged as a top regulator of adipose tissue phenotype with multiple binding sites in the leptin 3’-UTR that were confirmed with luciferase assay validations. miR-29 gain and loss-of-function modulated leptin mRNA and protein levels in primary adipocytes, and miR-29 abundance inversely correlated with leptin levels in adipose tissues. This work represents the only experimentally generated miRNA targetome in adipose tissue and identifies the first known post-transcriptional regulator of leptin.
O’Connor, Sean, "A Network Approach to Understanding miRNA Regulation in Adipose Tissue" (2020). Student Theses and Dissertations. 705.