BMP-dependent pathways and Alzheimer's disease

SUMMARY Amongst lipids that regulate intracellular transport routes, bis(monoacylglycerol)phosphate (BMP) is uniquely located to multivesicular bodies (MVBs) and late endosomes/lysosomes (LE/Lys), where it controls intra-lumenal vesicles (ILVs) formation. Upon fusion of MVBs with the membrane, ILVs will become exosomes. BMP thus regulates the correct addressing of cargoes to recycling, lysosomal degradation or extracellular exosomal export. Pathways regulated by BMP are profoundly altered in neurodegenerative diseases and levels of BMP are modified in these disorders -e.g., in human autopsy brain samples of late onset Alzheimer’s disease (LOAD). The ability to modulate BMP levels would open new perspectives in our understanding of the pathogenesis of these disorders and could provide new therapeutic options. The key obstacle to this approach is that the enzymes regulating BMP biosynthesis were so far unknown. Our lab recently identified an enzyme with phospholipase A2 (PLA2) activity -hereafter PLA2- regulating the limiting step of BMP synthesis in liposomes and in HeLa cell lines. Here we propose that modulating PLA2 levels and accompanying BMP levels could be an attractive strategy to modify pathological outcomes in LOAD. However, there is little that is known about PLA2 function and localization in the brain. In Aim 1, we will test the existence of a direct link between PLA2 protein levels and LOAD pathology in human autopsy brain samples. Specifically, we will use frozen samples from entorhinal cortex, hippocampus, prefrontal cortex and cerebellum of patients with severe or mild LOAD and compare them with unaffected control individuals. Using a combination of western blot and lipidomics, we will test (i) if protein levels of PLA2 are modified in pathology-prone brain regions of LOAD patients compared to controls, (ii) if protein levels of PLA2 correlate with disease progression and (iii) if BMP levels correlate with PLA2 levels. In Aim 2, we will gather evidence pertaining to the most disease-relevant brain cell type and biological outcomes to target in a future disease-modifying strategy. We will first identify the brain cell type localization of the PLA2 protein, using co-staining for PLA2 and different cell type markers in control human autopsy brain samples sections, in wild- type mouse brain sections and in microglia/astrocyte/neuron primary tri-cultures. In parallel, we will test whether modifying PLA2 protein levels leads to altered endolysosomal function and affects LOAD-relevant outcomes such as amyloid processing and exosomal release. We will down-regulate or overexpress Pla2 in mouse microglia/astrocyte/neuron primary tri-cultures and test whether it affects BMP levels, amyloid production and exosomal release, using a combination of confocal microscopy, western blot and Meso Scale Discovery (MSD) assays. If successful, the proposed studies will enable us to establish a strong baseline of experimental evidence on which we can build a robust disease-modifying strategy, to be investigated in a future larger application.