P3-441: Expression of mRNAs regulating local translation in synaptic terminals of MCI and early ad brain

Abstract

Recent evidence suggests Alzheimer's Disease (AD) begins with subtle alterations of synaptic function producing memory deficits prior to histopathological changes. Decreased expression of presynaptic proteins before synapse loss is part of a continuum of modifications during clinical progression of dementia. However, activity–dependant expression of proteins locally translated in both pre–and post–synaptic terminals, and mRNA binding proteins regulating their translation, have not been assessed in the AD brain. Perturbation of synaptic protein synthesis could result from modifications in structure and function of the pre–synaptic terminal and/or direct effects of Ab peptides at the post–synaptic terminal. To compare expression of mRNAs locally translated at the synapse in MCI/early AD versus controls. Patients were grouped primarily according to MMSE but also adjusted for amyloid burden. Frontal cortex (Area 9) was obtained post–mortem, from 8 age–matched controls, 6 MCI/early AD and 10 moderate–to–severe AD patients. Synaptoneurosomes were prepared and mRNA extracted for Affymetrix microarray profiling and quantitative PCR. Clustering of genes and fold change data was prepared with the Bioconductor package. Gene annotation and gene ontology groupings were identified using EASE (http://david.niaid.nih.gov/david/ease.htm). We screened mRNA sequences of changed genes for 3∼ UTR regulatory motifs, such as the cytoplasmic polyadenylation element (CPE), which regulate local translation at the synapse. In MCI there is up–regulation of genes involved in synaptic transmission, transport (including vesicle–mediated transport) and cell recognition, and down–regulation of cell–cycle genes, (p<0.01). Approximately 50% of up–regulated genes code for proteins localized to the membrane. We found 50 putative CPE–containing mRNAs out of 304 up–regulated (16%) and 22 of 157 down–regulated (14%). Genes encoding the mRNA–binding proteins pumilio homolog and HuD, and some corresponding mRNAs they regulate, were also up–regulated in MCI. Synaptoneurosome samples were evaluated for in vitro translation using biotin–labeled tRNA. Immunoblots revealed biotin labeled aCAMKII in all synaptosomal preparations tested. Thus, using human synaptoneurosomes, we have enriched a group of mRNAs with regulatory sequences involved in de novo protein synthesis at the synapse. Microarray profiling reveals that some of these genes are impacted in MCI brain and may be implicated in functional impairment during AD progression.