Abstract
The trafficking behavior of the lipid raft-dwelling US9 protein from Herpes Simplex Virus strikingly overlaps with that of the amyloid precursor protein (APP). Both US9 and APP processing machinery rely on their ability to shuttle between endosomes and plasma membranes, as well as on their lateral accumulation in lipid rafts. Therefore, repurposing US9 to track/modify these molecular events represents a valid approach to investigate pathological states including Alzheimer’s disease and HIV-associated neurocognitive disorders where APP misprocessing to amyloid beta formation has been observed. Accordingly, we investigated the cellular localization of US9-driven cargo in neurons and created a US9-driven functional assay based on the exogenous enzymatic activity of Tobacco Etch Virus Protease. Our results demonstrate that US9 can direct and control cleavage of recombinant proteins exposed on the luminal leaflet of transport vesicles. Furthermore, we confirmed that US9 is associated with lipid-rafts and can target functional enzymes to membrane microdomains where pathologic APP-processing is thought to occur. Overall, our results suggest strongly that US9 can serve as a molecular driver that targets functional cargos to the APP machinery and can be used as a tool to study the contribution of lipid rafts to neurodegenerative disease conditions where amyloidogenesis has been implicated.
Highlights
Metabolic processing of amyloid precursor protein (APP), a ubiquitously expressed protein, depends on cellular compartmentalization and lipid raft localization[12,13]
We exploited the properties of the Herpes Simplex Virus 1 (HSV-1) protein US9, which is enriched in lipid rafts, but has no toxic or catalytic activity
We designed a US9-driven functional assay based on the catalytic activity of an exogenous enzyme, the Tobacco Etch Virus (TEV) Protease, to test the hypothesis that US9 can target APP machinery compartmentalization and metabolism and as a proof of concept for further manipulation of APP processing in the presence of Human Immunodeficiency Virus (HIV) neurotoxins
Summary
Metabolic processing of APP, a ubiquitously expressed protein, depends on cellular compartmentalization and lipid raft localization[12,13]. To this end, we exploited the properties of the Herpes Simplex Virus 1 (HSV-1) protein US9, which is enriched in lipid rafts, but has no toxic or catalytic activity. As we recently demonstrated[32], the cellular distribution and functional properties of US9 do not require additional viral factors, are solely dependent on US9 sequence, and are not affected by attachment of a reporter/ cargo These US9 behaviors are maintained in multiple cell types, both primary and cultured lines, from different species[32]. The results indicate that US9 is able to drive functional cargos to lipid raft domains and is a powerful tool for use in characterizing lipid-raft dependent processes that span multiple cellular compartments, such as those that occur in metabolism of APP
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