Proteolytic cleavage of the low density lipoprotein (LDL) receptor: a novel regulator of plasma LDL cholesterol?

Cardiovascular disease, a term that generally refers to conditions that involve narrowed or blocked blood vessels that can lead to a heart attack, chest pain (angina) or stroke, kills one in three people in the UK. One of the most significant risk factors for cardiovascular disease is elevated levels of low-density lipoprotein (LDL) cholesterol - often referred to as 'bad cholesterol' - in the blood. The amount of LDL-cholesterol in the blood is controlled by its uptake into liver cells. On the surface of the liver cells is a specific protein, the LDL receptor, that binds the LDL-cholesterol. The number of active LDL receptors on the surface of liver cells is the single most important factor in regulating the amount of LDL-cholesterol in the blood. Through a series of molecular and cell-based studies we have identified that the LDL receptor is cut by the enzyme BMP1. This cleavage of the receptor by BMP1 reduces the ability of the receptor to take up the LDL-cholesterol into the cells. From these observations we hypothesise that proteolytic cleavage of the human LDL receptor by BMP1 regulates LDL-cholesterol uptake and hence the blood level of LDL-cholesterol. The overall aim of this project is to test the hypothesis that proteolytic cleavage of the human LDL receptor by BMP1 regulates LDL receptor function and hence plasma LDL cholesterol in vivo, and investigate the molecular and cellular mechanisms underpinning this. This will include utilizing mouse models to determine whether regulation of the activity of BMP1 may be a novel mechanism to lower the concentration of LDL-cholesterol in the blood. The findings from this research will be of great importance, both in terms of understanding the increasingly complex processes underlying the regulation of LDL-cholesterol, as well as potentially providing a new mechanism that could be a target for novel cholesterol-lowering drugs.

Cardiovascular disease is, and will be for the foreseeable future, the leading cause of death in the world. One of the most significant risk factors for cardiovascular disease is elevated levels of plasma low-density lipoprotein (LDL) cholesterol. The number of functional LDL receptors (LDLRs) expressed on the surface of hepatocytes is the primary determinant of plasma LDL-cholesterol levels. LDLR binds plasma LDL particles, mediating their endocytosis, and thereby reducing plasma cholesterol. Using recombinant proteins, hepatic cell lines and ex vivo tissue, we have observed that human LDLR is proteolytically cleaved in its extracellular ligand binding domain by the secreted zinc metalloprotease bone morphogenetic protein-1 (BMP1). Critically, cleavage of LDLR by BMP1 reduces the endocytosis of LDL-cholesterol in a cellular system. These observations form the basis of a novel mechanism for the post-translational regulation of LDLR. We hypothesise that proteolytic cleavage of human LDLR by BMP1 regulates LDL uptake and hence plasma LDL cholesterol in vivo. To test this hypothesis our specific objectives are to quantitate the reduction in binding and cellular uptake of LDL by the BMP1 truncated LDLR; determine the effect of BMP1 regulation on LDLR proteolysis and plasma LDL cholesterol in vivo through complementary approaches of pharmacological inhibition and genetic modulation of BMP1 in mice expressing human LDLR; and understand the cellular and molecular mechanism of action of BMP1 on LDLR with particular emphasis on determining the subcellular compartment where BMP1 cleaves LDLR and whether BMP1 interacts with LDLR via an exosite. The results of this work will validate BMP1 as a novel regulator of human LDLR function, contribute to our understanding of the increasingly complex processes underlying the regulation of LDLR and plasma LDL cholesterol, as well as potentially identifying a new mechanism that could be a target for novel cholesterol-lowering drugs.

This research proposal relates to the advancement of fundamental science underpinning cholesterol metabolism. In the lifetime of this proposal, our research will impact upon academic researchers in a range of disciplines (from basic cell biology of LDLR through to medicinal chemists seeking to develop novel drugs to lower plasma cholesterol) both in the UK and internationally. Our work will add new knowledge and scientific advancement to our understanding of the post-translational regulation of LDLR and cholesterol metabolism. These will be achieved during the short to medium term. The other main beneficiaries of this proposal will be the PDRA, as this research provides an excellent opportunity for development of specialist scientific skills as well as transferrable skills, equipping the individual for a career in science or other fields.

In the longer term our work has the potential to deliver impact by benefitting the general public and the health services through identifying a novel therapeutic approach to lower plasma cholesterol which could have a global impact. If this is realised in the future, then our research could have the potential for enhancing the quality of life, health and well-being of significant numbers of individuals affected by high levels of LDL cholesterol, and also the research could deliver commercial impact in terms of development and commercialisation of novel therapeutic treatments, contributing to creation of economic growth and new employment opportunities. Benefits would also be delivered to Healthcare services and Healthcare professionals. Further economic and societal impact will be achieved through attracting research and development investment from pharmaceutical companies to exploit and develop our observations.