Proteomic Assessment of Post-Translational Modifications in the Brain of 5x-FAD Females

Abstract

Dementia prevalence is expected to more than double in the next three decades stressing numerous healthcare systems and caregivers across the globe. Alzheimer’s disease (AD), the most common form of dementia, is a multi-factorial neurodegenerative disease without a cure due to the incomplete understanding of its complex progression. Our aim is to report hypothesis generating data for targeted functional studies to understand the mechanisms underlying differential expression and post-translational modifications in AD using the 5x-FAD murine model. The bulk proteome, S-nitrosylated proteome, phosphoproteome, and acetylome were assessed by unbiased tandem mass spectrometry and label-free quantitative proteomics. Affinity purification assays were utilized to enrich samples to evaluate S-nitrosylated proteins. Western blots analysis was performed on specific protein targets of interest from the bulk proteome and S-nitrosylated proteome to validate proteomic results. We observed 426 proteins with differential expression patterns across the whole proteome and 225 proteins with alterations in S-nitrosylation in 6-monthold female 5x-FAD mice. Additionally, 161 phosphopeptides and 23 acetylated peptides were found to be significantly different compared to wild-type (WT) littermates. Gene ontology (GO) analysis implicated enrichment of glutamateric signaling, oxidoreudctase activity, and cytoskeletal proteins, supporting current AD literature, while also elucidating the novel targeting that post translational modifications could have on the electron transport chain, transcription, and histone and mitochondrial acetylation. We further show primary validation of increased glial fibrillary acidic protein expression patterns, known to occur in AD patients, and revealed a conserved decrease in S-nitrosylation of myelin proteolipid protein. However, myelin-associated oligodendrocyte basic protein quantification did not allow for conclusive inferences to an under investigated aspect of AD. These proteomic data concur with well-documented protein changes, post-translational modifications, and enriched signaling pathways found in humans living with AD and animal models of the disease, linking specific proteins involved with cytoskeletal organization, mitochondrial ATP genesis, ion homeostasis, transcription, translation, and myelin sheath maintenance contributing to the multifactorial nature of AD.