The Influence of APOEε4 on the pTau Interactome in Sporadic Alzheimer’s Disease

Background

Alzheimer’s disease (AD) is a neurodegenerative disease characterized by extracellular deposition and aggregation of β-amyloid protein (Aβ) forming various types of Aβ deposits and the intracellular accumulation and assembly of abnormally phosphorylated tau protein (p-tau) forming neurofibrillary tangles. The pathogenesis of AD is also closely linked to the neuroanatomical progression of tau pathology. These pathological markers sequentially affect different brain regions, following the Thal five stages (Aβ pathology) and Braak six stages (tau pathology).

Among the genetic risk factors for AD, the polymorphism of the Apolipoprotein E gene (APOE) is a major contributor. The APOEε4 allele is positively correlated with increased risk and earlier onset age of sporadic AD, while the APOEε2 allele is considered protective. APOE is a glycoprotein predominantly secreted by astrocytes, typically involved in the transport of phospholipids and cholesterol. It is known that APOE co-localizes with Aβ deposits, and the ε4 allele is closely associated with exacerbated Aβ pathology.

Although APOE has been extensively studied, its role in tau pathology remains unclear. Recent data suggest that APOE polymorphism also affects phosphorylated tau pathology. Some studies indicate that in vivo, ApoEε4 might affect p-tau accumulation independently of Aβ. However, the relationship between APOE and p-tau remains vague, requiring further research to elucidate APOE’s role in tau pathology.

Source of Study

This paper, titled “The Influence of APOEε4 on the pTau Interactome in Sporadic Alzheimer’s Disease,” is authored by Manon Thierry, Jackeline Ponce, Mitchell Martà-Ariza, among others. The research was conducted at multiple institutions, including the Departments of Neurology and Biochemistry and Molecular Pharmacology at NYU School of Medicine. The paper was published in the journal “Acta Neuropathologica” in 2024.

Research Methods

To reveal the influence of APOEε4 on p-tau, the research team adopted a series of experimental procedures, including anti-p-tau antibody immunoprecipitation and mass spectrometry. The specific process is as follows:

Research Process

Sample Collection and Processing: - Collected prefrontal cortex tissue from 25 sporadic AD patients from the Alzheimer’s Disease Research Centers at NYU and Columbia University. - Samples were grouped based on APOE genotype: APOEε3/ε3 group and APOEε4/ε4 group, matched for age, sex, and comorbidity status. - Samples were initially processed using standard biochemical and genotyping methods, including DNA extraction, PCR, and Sanger sequencing.

Immunoprecipitation and Mass Spectrometry Analysis: - Immunoprecipitation with anti-p-tau antibody PHF1, collecting p-tau and its binding partner proteins. - Following degradation, extraction, and elution, protein analysis was conducted via liquid chromatography-tandem mass spectrometry (LC-MS/MS). - Mass spectrometry data were processed using Proteome Discoverer software and the “Significance Analysis of Interactome Express Algorithm (SAINT)” to screen potential p-tau interacting proteins.

Biochemical Analysis and Validation: - Verified the enrichment of p-tau in immunoprecipitated products using Western Blot and silver staining techniques. - Combined with immunohistochemistry (IHC) analysis, further determined the impact of APOE genotype on p-tau subcellular localization and pathological characteristics.

Data Analysis and Network Functional Enrichment

To analyze the mass spectrometry data, the research team used tools like STRING and Cytoscape. Through Gene Ontology (GO) term analysis and network functional enrichment analysis, p-tau interacting proteins were identified and classified, confirming their roles in cellular components and biological processes.

Research Results

The research shows that the APOE genotype significantly affects the p-tau interactome. Major findings include:

1. Shared p-tau Interaction Proteins:

33 proteins were found to interact with p-tau, primarily related to the proteasomal system, involving mRNA metabolism and protein degradation processes.

2. p-tau Interaction Proteins Specific to APOEε3/ε3 and APOEε4/ε4:

• APOEε3/ε3: 47 p-tau interacting proteins, mainly nuclear proteins involved in RNA metabolism.
• APOEε4/ε4: 35 p-tau interacting proteins, mostly synapse-related proteins involved in cellular transport.

3. Tau Pathology Morphological Characteristics:

Immunohistochemistry analysis shows that APOEε4/ε4 group has a significantly higher density of p-tau positive neuronal nuclei compared to APOEε3/ε3 group, especially in axonal endings and synaptic regions, indicating that APOEε4 may promote the spread of tau pathology to brain regions affected by Aβ.

4. Cerebral Amyloid Angiopathy (CAA):

The APOEε4/ε4 group had higher frequency and severity of CAA compared to the APOEε3/ε3 group, indicating that APOEε4 is associated with exacerbation of CAA-related pathology.

Conclusion and Significance

The study demonstrates that the APOEε4 allele impacts AD risk not only by promoting Aβ pathology but also by significantly affecting the subcellular localization and interaction of p-tau, influencing the development of tau pathology. This finding provides key insights for identifying new therapeutic targets, particularly for treatment strategies targeting APOEε4 carriers.

Highlights and Novelty of the Study

• This study is the first to compare the p-tau interactome in sporadic AD between APOEε3/ε3 and APOEε4/ε4 carriers.
• It reveals that APOEε4 promotes the localization of p-tau at synapses and neuronal endings, contributing to the understanding of APOEε4’s role in promoting tau pathology progression.
• The data support that APOEε4 exacerbates AD pathology by affecting cellular transport and RNA metabolism processes.

Important Implications and Application Value

This study uncovers a novel mechanism of how APOEε4 influences AD pathology beyond Aβ pathology, emphasizing the significance of APOE genotype in personalized and targeted therapeutic strategies. Future research should further explore the interaction networks of different tau epitopes across various AD stages to provide a theoretical basis for more precise early diagnosis and intervention.