Bifunctional Anti-Non-Amyloid Component α-Synuclein Nanobodies Are Protective In Situ

Misfolding, abnormal accumulation, and secretion of α-Synuclein (α-Syn) are closely associated with synucleinopathies, including Parkinson's disease (PD). VH14 is a human single domain intrabody selected against the non-amyloid component (NAC) hydrophobic interaction region of α-Syn, which is c...

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Bibliographic Details
Published in:PloS one 2016-11, Vol.11 (11), p.e0165964-e0165964
Main Authors: Butler, David C, Joshi, Shubhada N, Genst, Erwin De, Baghel, Ankit S, Dobson, Christopher M, Messer, Anne
Format: Article
Language:English
Subjects:
alpha-Synuclein - chemistry
alpha-Synuclein - immunology
alpha-Synuclein - physiology
Amyloidogenic Proteins - immunology
Animals
Annexin V
Antigens
Apoptosis
Autophagy
Binding sites
Biocompatibility
Biology and Life Sciences
Blotting, Western
C-Terminus
Cell culture
Cell Line
Cell lines
Cloning
Deoxyribonucleic acid
Disease
DNA
Drosophila
Flow Cytometry
Humans
Huntingtin
Hydrophobicity
Immunoglobulins
Insects
Iodides
Medicine and Health Sciences
Movement disorders
Nanobodies
Neurodegenerative diseases
Neurons - metabolism
Neurosciences
Parkinson's disease
Plasmids
Propidium iodide
Proteasome Endopeptidase Complex - metabolism
Proteasomes
Proteins
Rats
Research and Analysis Methods
Secretion
Single-Domain Antibodies
Stem Cells
Synuclein
Toxicity
Transfection
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Description
Summary:Misfolding, abnormal accumulation, and secretion of α-Synuclein (α-Syn) are closely associated with synucleinopathies, including Parkinson's disease (PD). VH14 is a human single domain intrabody selected against the non-amyloid component (NAC) hydrophobic interaction region of α-Syn, which is critical for initial aggregation. Using neuronal cell lines, we show that as a bifunctional nanobody fused to a proteasome targeting signal, VH14PEST can counteract heterologous proteostatic effects of mutant α-Syn on mutant huntingtin Exon1 and protect against α-Syn toxicity using propidium iodide or Annexin V readouts. We compared this anti-NAC candidate to NbSyn87, which binds to the C-terminus of α-Syn. NbSyn87PEST degrades α-Syn as well or better than VH14PEST. However, while both candidates reduced toxicity, VH14PEST appears more effective in both proteostatic stress and toxicity assays. These results show that the approach of reducing intracellular monomeric targets with novel antibody engineering technology should allow in vivo modulation of proteostatic pathologies.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0165964