Amyloid-β Peptides Disrupt Interactions Between VAMP-2 and SNAP-25 in Neuronal Cells as Determined by FRET/FLIM.
BACKGROUND
Synaptic dysfunction prevalent in Alzheimer's disease (AD) brain is closely associated with increased accumulation of amyloid-β (Aβ) peptides in the brain parenchyma. It is widely believed that Aβ peptides trigger synaptic dysfunction by interfering with the synaptic vesicular fusion and the release of neurotransmitters, primarily facilitated by the SNARE protein complexes formed by VAMP-2, SNAP-25, and syntaxin-1. However, Aβ interactions with SNARE proteins to ultimately disrupt synaptic vesicular fusion are not well understood.


OBJECTIVE
Our objective is to elucidate mechanisms by which Aβ peptides perturb SNARE complexes.


METHODS
Intensity (qualitative) and lifetime (quantitative) based measurements involving Forster (fluorescence) resonance energy transfer (FRET) followed by fluorescence lifetime imaging microscopy (FLIM) were employed to investigate the effect of Aβ peptides on dynamic interactions between VAMP-2, labeled with cerulean (Cer) at the N-terminus (FRET donor), and SNAP-25 labeled with citrine (Cit) on the N-terminus (FRET acceptor). The FRET and FLIM interactions at the exocytosis locations on the pre-synaptic membrane were recorded under spontaneous and high potassium evoked conditions. Moreover, cellular accumulation of fluorescein labeled Aβ (F-Aβ) peptides and their co-localization with Cer-VAMP2 was investigated by confocal microscopy.


RESULTS
The F-Aβ40 and F-Aβ42 are internalized by differentiated N2A cells, where they colocalize with Cer-VAMP2. Both Aβ40 and Aβ42 decrease interactions between the N-termini of Cer-VAMP2 and Cit-SNAP25 in N2A cells, as determined by FRET/FLIM.


CONCLUSION
By perturbing the N-terminal interactions between VAMP-2 and SNAP-25, Aβ40 and Aβ42, can directly interfere with the SNARE complex formation, which is critical for the docking and fusion of synaptic vesicles.
['Sharda|Nidhi|N|', 'Pengo|Thomas|T|', 'Wang|Zengtao|Z|', 'Kandimalla|Karunya K|KK|']
D016229:Amyloid beta-Peptides D045744:Cell Line, Tumor D031541:Fluorescence Resonance Energy Transfer D006801:Humans D008856:Microscopy, Fluorescence D009474:Neurons D011485:Protein Binding D050825:Synaptosomal-Associated Protein 25 D050988:Vesicle-Associated Membrane Protein 2
Amyloid-β peptides VAMP-2 SNAP-25 FRET FLIM synaptic vesicular fusion SNAREs exocytosis Alzheimer’s disease