There is currently little known about the role of extracellular matrix (ECM) in the regulation of microRNA (miRs), a family of short, non-coding RNA that repress gene expression at the post-translational level by binding to the 3'-untranslated region (3'UTR) of target mRNA.This thesis uses the mouse mammary gland (MG) to address this question by investigating whether the extracellular matrix regulates miR expression in mammary epithelial cells (MECs).miR expression profiles were generated using MECs cultured in 2D on collagen Ι and in 3D on laminin-rich basement membrane (LrBM). I identified 88 miRs that are more highly expressed in collagen cultured MECs and 8 miRs that have higher expression in MECs cultured on LrBM including miR-146b, a miR known to reduce metastases to the lung in breast cancer. The culture model used compares not only collagen to LrBM but also a stiff environment to a soft environment; raising the question of whether miR-146b is regulated by MEC interaction with ECM proteins or by cellular tension imposed by the microenvironment.Further investigation into miR-146b expression in MECs showed that its expression also increases in response to prolactin stimulation. Expression of the prolactin receptor and subsequently prolactin signalling is reduced in MECs cultured on collagen, but increases in MECs treated with blebbistatin or Y27632, which release cellular tension. However, neither drug had any affect on expression of miR-146. The ECM adhesion receptor β1-intregrin regulates MEC differentiation via cross-talk with prolactin receptor signalling. By using MECs from β1- itgfx/fx;CreER mice I identified a novel mechanism of miR regulation in which β1-intregrin signalling regulates transcription of miR-146b.This study has shows the importance of ECM in the regulation of miR expression and, whilst further investigations are still required, highlights the importance of ECM culture models in studying miR expression and function.
|Date of Award||1 Aug 2013|
- The University of Manchester
|Supervisor||Charles Streuli (Supervisor)|