Differential expression and redox proteomics analyses of an Alzheimer disease transgenic mouse model: effects of the amyloid-β peptide of amyloid precursor proteinΞ

Abstract

Among the pathological factors known to be associated with Alzheimer disease (AD), oxidative stress induced by the amyloid-β peptide (Aβ) has been demonstrated to play a key role in human brain and animal models of AD. Recently, we reported elevated levels of oxidative damage in the brain of a transgenic (Tg) AD mouse model with Swedish and Indiana familial AD mutations in human amyloid precursor protein (APP) [PDAPP mice, line J20], as evidenced by increased levels of protein carbonyls, 3-nitrotyrosine, and protein-bound 4-hydroxy-2-nonenal. This oxidative damage was dependent on the methionine 35 residue within the Aβ peptide. Further insight into the molecular pathways affected in this Tg model of AD may be gained with discovery-based proteomics studies; therefore, two-dimensional gel-based expression proteomics was performed to compare differences in brain protein levels of J20 Tg mice with non-transgenic (NTg) littermate controls. Based on our studies, we identified six proteins that had significantly increased levels in J20 Tg relative to NTg mice: calcineurin subunit B type 1, ρ GDP-dissociation inhibitor 1, Tcomplex protein 1 subunit α A, α-enolase, peptidyl-prolyl cis-trans isomerase (Pin-1), and ATP synthase subunit α mitochondrial. Several of these proteins have previously been implicated in in vitro and in vivo models and subjects with AD. Additionally, using redox proteomics analyses we identified two oxidatively-modified proteins: phosphatidylethanolamine-binding protein 1 and Pin-1 with decreased levels of protein 3-nitrotyrosine in J20 Tg mice relative to NTg. Western blotting and immunoprecipitation analyses were used to validate proteomics results. Overall, these Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Lahad et al., 1995, Slooter et al., 1998, endothelial nitric oxide synthase-3 (Dahiyat et al., 1999), and α2-macroglobulin (Blacker et al., 1998) have been found in other AD cases. Histopathological hallmarks of AD include senile plaques (SP), neurofibrillary tangles (NFT), and synapse loss. Additionally, oxidative stress has been implicated in the pathogenesis of AD (Smith et al., 1994, Good et al., 1996, Markesbery, 1997, Smith et al., 1997, Butterfield and Lauderback, 2002). NIH Public AccessSPs are largely composed of amyloid-β (Aβ) peptides, which are generated by β-and γ-secretase cleavage of the APP protein.The most common forms of Aβ associated with human AD are Aβ(1-40) and Aβ(1-42) (Selkoe, 1996), and the latter has been shown to be more toxic than Aβ(1-40) in model systems of AD , Boyd-Kimball et al., 2005a, 2005b, 2005c, Mohmm...

Do Proteomics Analyses Provide Insights into Reduced Oxidative Stress in the Brain of an Alzheimer Disease Transgenic Mouse Model with an M631L Amyloid Precursor Protein Substitution and Thereby the Importance of Amyloid-Beta-Resident Methionine 35 in Alzheimer Disease Pathogenesis?

“…The increase in the levels of mitochondrial-related proteins such as ATP synthase subunit a, VATB2, and VDAC1 could indicate that their levels are decreased in J20 Tg mice due to the presence of Met in Ab, while M631L Tg mice are unaffected. In our previous studies conducted on the proteome changes in J20 Tg mice, we observed increased expression levels of glycolytic and mitochondrial proteins (6). VATB2 also plays a role in autophagy, which is consistent with the suggestion that in the absence of oxidative stress due to the absence of Met-35 in Ab(1-42) of APP, autophagic processes work well, but are compromised in J20 mice.…”

“…In the current study, we observed decreased levels of Pin-1 in M631L Tg mice relative to J20 Tg mice. In the comparison between J20 Tg mice relative to J20 NTg animals, we previously observed an increase in Pin-1 expression, which may indicate the brain's attempt to reduce elevated Ab levels and prevent oligomeric Ab formation (6).…”

“…Figure 3 shows representative 2D-gel images of protein spots observed to statistically change in density between M631L Tg and J20 Tg mice. The J20 Tg data were recently published in our initial proteome investigations in this mouse model (6), and are used here to allow direct proteome comparisons based on the presence or absence of Met-35 in the Ab(1-42) portion of APP. From these comparisons and MS/MS analyses of tryptic digest solutions of individual spots, 14 differentially expressed proteins were identified (Table 2).…”

“…All mice were 9 months of age at sacrifice. We note that the J20 Tg mice and corresponding 2D-gels used in these studies were used in our recent report of proteome alterations in J20 Tg mice (6).…”

Protein Thiol Modification and Thiol Proteomics

“…Redox proteomic analysis of the post-mortem brains of AD patients revealed the presence of oxidative modifications of various protein substrates. Among them, some are relevant mitochondrial proteins, such as ATP synthase -and -chain and VDAC (Robinson et al, 2011). These mitochondrial r e s i d e n t p r o t e i n s w e r e f o u n d t o b e o x i d ised in the hippocampus and the observed modifications could play a role in the mitochondrial dysfunction and cell death observed in AD.…”