A genomic, proteomic and transcriptomic investigation into growth hormone actions

Abstract

Interactions between genomic loci have been implicated in playing a critical role in the regulation of gene transcription. Growth hormone (GH) is a peptide hormone predominantly produced in the pituitary that is crucial for normal growth and metabolism. GH actions are activated through binding to its cell-surface receptor, the GH receptor (GHR), with consequent activation of downstream signalling cascades. Genetic variation in the GH locus genes and aberrant GH signalling have been implicated in diseases such as cancer. This led to the hypothesis that genetic variation that occurs in the GH locus may potentially result in alteration of gene regulatory mechanisms. Common SNPs in the regions across the GH locus that physically interacted with genes, both in cis and trans (intra and inter-chromosomal), were identified. These SNPs were also found to be associated with altered expression of these genes. This suggests that regions encompassing the GH locus function as regulatory hubs for multiple genes, some of which are involved in cellular and cancer pathways, related to GH/GHR signalling. Nuclear localisation of GHR and its association with increased tumorigenesis has previously been reported, but the specific consequences of this phenomenon remain unclear. To determine whether there is a functional role for the GHR in the nucleus, combined immunoprecipitation-mass spectrometry was used to assess whether the GHR interacts with proteins in the nucleus and microarray analysis was used to determine the consequent effects of this nuclear import on gene expression. Multiple proteins were found to localise with the GHR, including two transcription factors, HMGN1 and SUMO1. Moreover, targets genes of HMGN1 and SUMO1 were found to be differentially expressed following GH treatment. This suggests that GHR nuclear translocation potentially serves as an auxiliary mechanism for regulation of GH actions. The competitive endogenous RNA (ceRNA) theory suggests that miRNA can be sequestered by lncRNA through competition for shared binding sites, causing changes in miRNA targeted gene expression. GH/GHR signalling has been linked to altered non-coding RNA transcription profiles. As such, a GH-induced ceRNA network was constructed to elucidate the molecular regulatory mechanisms involved in mediating GH functions in a mammary epithelial cell line. This thesis provides novel insights into the mechanisms underlying GH actions, including a functional role for the GHR in the nucleus and the coordinated regulation of endogenous RNA networks.