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PATHOLOGICAL OLIGOMERISATION OF PHOSPHORYLATED NUCLEAR ASYN IS ASSOCIATED WITH ELEVATED DNA DAMAGE IN DEMENTIA WITH LEWY BODIES.
Abstract
Aims
Dementia with Lewy bodies (DLB) is defined by the cytoplasmic accumulation of alpha-synuclein (aSyn) within neurons. Although predominately pre-synaptic, nuclear aSyn (aSynNuc) has been frequently observed in-vitro where it is associated with altered DNA integrity. Yet the presence and role of aSynNuc in the human brain remains controversial and its relevance to DLB pathology unclear, here we set out to resolve the existence and relevance of aSynNuc.
Methods
aSynNuc was investigated via immunohistochemical analysis of fixed postmortem brain tissue and in isolated nuclear preparations from frozen brain tissue. Nuclear preparations were additionally subject to western-blot and label-free mass spectrometry.
Results
Immunohistochemical analysis demonstrated intra-nuclear puncta reactive to a variety of pan-aSyn antibodies as well as to the disease associated phosphorylation specific pS129-aSyn antibody. The presence of aSynNuc was further confirmed via mass spectrometry and biochemical analysis. In all cases, monomeric aSynNuc was detected at ~10 fold lower levels compared to cytoplasmic aSyn. Critically, only in DLB cases were aSynNuc oligomers evident, with both monomers and oligomers being highly phosphorylated. Pathological aSynNuc was observed alongside elevated levels of DNA double strand breaks (DSBs). Elevated DNA damage was observed in the absence of changes to upstream DNA damage signalling kinases (ATM/ATR) suggesting impairments in DSB repair as opposed to an increase in the occurrence of DSB.
Conclusions
Our data support emerging roles for aSyn in DNA damage repair and highlights a novel disease mechanism underlying the neurodegenerative processes of DLB and implicates genomic homeostasis as a potential future therapeutic target.