Cytosolic proteins lose solubility as amyloid deposits in a transgenic mouse model of Alzheimer-type amyloidosis

Abstract

The extracellular accumulation of β-amyloid peptide is a key trigger in the pathogenesis of Alzheimer's disease (AD). In humans, amyloid deposition precedes the appearance of intracellular inclusion pathology formed by cytosolic proteins such as Tau, α-synuclein and TDP-43. These secondary pathologies have not been observed in mice that model Alzheimer-type amyloidosis by expressing mutant amyloid precursor protein, with or without mutant presenilin 1. The lack of secondary pathology in these models has made it difficult to establish how amyloid deposition initiates the cascade of events that leads to secondary intracellular pathology that characterizes human AD. In transgenic mice that model Alzheimer-type amyloidosis, we sought to determine whether there is evidence of altered cytosolic protein folding by assessing whether amyloid deposition causes normally soluble proteins to misfold. Using a method that involved detergent extraction and sedimentation coupled with proteomic approaches, we identified numerous cytosolic proteins that show specific losses in solubility as amyloid accumulates. The proteins identified included glycolytic enzymes and members of the 14-3-3 chaperone family. A substantial accumulation of lysine 48-linked polyubiquitin was also detected. Overall, the data demonstrate that the accumulation of amyloid by some manner causes the loss of solubility intracellular cytosolic proteins.

Non-targeted Identification of Prions and Amyloid-forming Proteins from Yeast and Mammalian Cells

“…Although other mass spectrometry-based approaches were employed previously to characterize proteins associated with amyloid (Ref. 62 and references therein), our method is unique as it includes both stringent SDS treatment and a novel separation of aggregates. As a consequence, only the strongest interactors survived, reducing the burden of analyzing hundreds of hits.…”

The Biology of Proteostasis in Aging and Disease

“…This reduction in chaperone levels can occur through a combination of transcriptional dysregulation and sequestration of chaperones by mHTT or polyQ aggregates (64–66). Sequestration of chaperones by insoluble aggregates also leads to reduced levels of soluble HSP70, HSP40, HSP90, and sHSPs in mouse and C. elegans models of AD, tissue culture and Drosophila models of PD (67–70), and tissue culture and mouse models of spinocerebellar ataxia 3 [SCA3; also known as Machado–Joseph disease (MJD)] (71). These observations suggest that perturbation of molecular chaperones may be central to global proteostasis collapse in disease.…”

Vulnerability of newly synthesized proteins to proteostasis stress

“…To determine which types of proteins lose solubility in these HEK293 cells, we analyzed the LC-MS/MS data using a statistical method as previously described (Xu et al, 2012(Xu et al, , 2013. Apart from ubiquitin, we identified 13 additional proteins in which the frequency of peptide identification in the DOC-insoluble fraction of heat-shocked cells was significantly (P<0.05) more abundant than predicted in two experiments (Table 1).…”