Apolipoprotein E (ApoE) is the strongest genetic risk factor for late-onset Alzheimer?s disease (AD). Individuals who possess one or more copies of the APOE4 variant have an increased risk of developing AD, with an earlier age of onset and more rapid cognitive decline, compared to those with the more common APOE3 allele. ApoE is expressed primarily in astrocytes, and secreted on lipoprotein particles to provide cholesterol and unsaturated phospholipids for membrane expansion and synapse formation in neurons. However, little is known regarding the basic molecular mechanisms behind ApoE function in astrocytes, and how the E4 variant promotes disease. I have discovered that ApoE can traffic either to the secretory pathway or to cytoplasmic lipid droplets in astrocytes. My central hypothesis is that ApoE can be stimulated to target lipid droplets in the cytoplasm rather than being secreted, and that it is involved in promoting the storage of neutral lipids within lipid droplets. I also hypothesize that the lipid droplet function of ApoE is altered in the disease-associated E4 and E2 variants. To test this hypothesis, I will perform structure-function studies to determine which regions of the protein are necessary for ApoE to target the cytoplasmic compartment and localize to lipid droplets. I will examine how ApoE trafficking is controlled by neuronal signals by performing astrocyte-neuron coculture assays. To determine the function of ApoE targeted to lipid droplets, I will use shRNA to knockdown ApoE and observe the effect on the distribution of neutral lipids in the cell. I will then determine whether ApoE regulates lipid droplet biogenesis, growth, or turnover via fluorescent lipid pulse-chase assays coupled with time-lapse microscopy and automated image analysis. I will compare the trafficking and function of the different variants of ApoE to test whether the lipid droplet function is altered in E2 and E4. This project will uncover how ApoE variants regulate lipid droplet metabolism in astrocytes.
Apolipoprotein E (ApoE) is the major genetic risk factor for sporadic, late-onset Alzheimer?s disease, yet little is known about the function of ApoE in astrocytes. We have discovered that in addition to being secreted, ApoE can traffic to cytoplasmic lipid droplets in astrocytes. Understanding the role of ApoE on astrocyte lipid droplet metabolism, and how this function is altered in disease-associated variants, will improve our understanding behind the molecular mechanisms that predispose individuals to develop Alzheimer?s disease.
