ApoE lipidation, not isoform, is the key modulator of Aβ interaction, uptake, and cytotoxicity
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Background
Inherited variations in the Apolipoprotein E ( APOE ) gene are the largest genetic determinant for late-onset Alzheimer’s Disease, with the APOEε4 allele conferring the highest risk. While APOE was shown to modulate amyloid beta (Aβ) pathology in a genotype-specific manner ( APOEε4 > APOEε3 > APOEε2 ), it remains an important open question whether these differences are due directly to isoform-specific interactions between ApoE and Aβ or indirect effects on Aβ clearance. This is further complicated because ApoE exists in lipidated or unlipidated states, which influence its biochemical properties. To disentangle how single ApoE mutations confer vastly different effects on Aβ pathology, we investigated how both the ApoE isoform and lipidation modulate its interaction with inert and cytotoxic Aβ species, and its effect on Aβ uptake and cytotoxicity in human astrocytes.
Methods
We prepared Aβ in distinct aggregation states (monomers, oligomers, and fibrils) and ApoE in different lipidation states to characterize their size and affinity for one another using fluorescence correlation spectroscopy and fluorescence polarization, respectively. We then utilized flow cytometry and live cell imaging to quantify the uptake of Aβ in different aggregation states by primary and immortalized human astrocytes in the presence of different isoforms of lipidated or unlipidated ApoE.
Results
This study revealed that ApoE lipidation, not isoform, has the biggest impact on its interaction with Aβ, on the uptake of Aβ by astrocytes, and on Aβ-induced cytotoxicity. Specifically, unlipidated ApoE preferentially interacts with Aβ oligomers and fibrils, which substantially inhibits their uptake by astrocytes. Conversely, lipidated ApoE showed no interaction with Aβ oligomers and had a reduced ability to inhibit Aβ uptake.
Conclusions
Our observations provide important molecular details that suggest previously observed ApoE isoform-specific differences in AD risk are likely driven primarily by in vivo differences in ApoE lipidation, rather than biophysical differences between ApoE isoforms in their interaction with Aβ. We propose that the impaired lipidation of ApoE4 in the brain increases levels of unlipidated ApoE, which then bind toxic Aβ oligomers and reduce their clearance by astrocytes. These findings underscore the therapeutic potential of interventions aimed at increasing ApoE lipidation and decreasing interactions between toxic Aβ oligomers and ApoE.
