Dual-tagged amyloid-β precursor protein reveals distinct transport pathways of its N- and C-terminal fragments

Christine Villegas, Virgil Muresan, Zoia Ladescu Muresan

Research output: Contribution to journalArticlepeer-review

10 Scopus citations

Abstract

The amyloid-β precursor protein (APP), a type I transmembrane protein genetically associated with Alzheimer's disease, has a complex biology that includes proteolytic processing into potentially toxic fragments, extensive trafficking and multiple, yet poorly-defined functions. We recently proposed that a significant fraction of APP is proteolytically cleaved in the neuronal soma into N- and C-terminal fragments (NTFs and CTFs), which then target independently of each other to separate destinations in the cell. Here, we prove this concept with live imaging and immunolocalization of two dual, N- and C-termini-tagged APP constructs: CFP-APP-YFP [containing the fluorescent tags, cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP)] and FLAG-APP-Myc. When expressed at low levels in neuronal cells, these constructs are processed into differently tagged NTFs and CTFs that reveal distinct distributions and characteristics of transport. Like the endogenous N- and C-terminal epitopes of APP, the FLAG-tagged NTFs are present in trains of vesicles and tubules that localize to short filaments, which often immunostain for acetylated tubulin, whereas the Myc-tagged CTFs are detected on randomly distributed vesicle-like structures. The experimental treatments that selectively destabilize the acetylated microtubules abrogate the distribution of NTFs along filaments, without altering the random distribution of CTFs. These results indicate that the NTFs and CTFs are recruited to distinct transport pathways and reach separate destinations in neurons, where they likely accomplish functions independent of the parental, full-length APP. They also point to a compartment associated with acetylated microtubules in the neuronal soma-not the neurite terminals-as a major site of APP cleavage, and segregation of NTFs from CTFs.

Original languageEnglish (US)
Article numberddt555
Pages (from-to)1631-1643
Number of pages13
JournalHuman molecular genetics
Volume23
Issue number6
DOIs
StatePublished - Mar 2014

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics
  • Genetics(clinical)

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