Alpha-synuclein is a presynaptic protein that forms abnormal cytoplasmic aggregates in Lewy body disorders. Although nuclear alpha-synuclein localization has been described, its function in the nucleus is not well understood. We demonstrate that alpha-synuclein modulates DNA repair. First, alpha-synuclein colocalizes with DNA damage response components within discrete foci in human cells and mouse brain. Removal of alpha-synuclein in human cells leads to increased DNA double-strand break (DSB) levels after bleomycin treatment and a reduced ability to repair these DSBs. Similarly, alpha-synuclein knock-out mice show increased neuronal DSBs that can be rescued by transgenic reintroduction of human alpha-synuclein. Alpha-synuclein binds double-stranded DNA and helps to facilitate the non-homologous end-joining reaction. Using a new, in vivo imaging approach that we developed, we find that serine-129-phosphorylated alpha-synuclein is rapidly recruited to DNA damage sites in living mouse cortex. We find that Lewy inclusion-containing neurons in both mouse model and human-derived patient tissue demonstrate increased DSB levels. Based on these data, we propose a model whereby cytoplasmic aggregation of alpha-synuclein reduces its nuclear levels, increases DSBs, and may contribute to programmed cell death via nuclear loss-of-function. This model could inform development of new treatments for Lewy body disorders by targeting alpha-synuclein-mediated DNA repair mechanisms.

Phosphorylation of alpha-synuclein at serine-129 is an important marker of pathologically relevant, aggregated forms of the protein in several important human diseases, including Parkinson’s disease, Dementia with Lewy bodies, and Multiple system atrophy. Although several kinases have been shown to be capable of phosphorylating alpha-synuclein in various model systems, the identity of the kinase that phosphorylates alpha-synuclein in the Lewy body remains unknown. One member of the Polo-like kinase family, PLK2, is a strong candidate for being the Lewy body kinase. To examine this possibility, we have used a combination of approaches, including biochemical, immunohistochemical, and in vivo multiphoton imaging techniques to study the consequences of PLK2 genetic deletion on alpha-synuclein phosphorylation in both the presynaptic terminal and preformed fibril-induced Lewy body pathology in mouse cortex. We find that PLK2 deletion reduces presynaptic terminal alpha-synuclein serine-129 phosphorylation, but has no effect on Lewy body phosphorylation levels. Serine-129 mutation to the phosphomimetic alanine or the unphosphorylatable analog aspartate does not change the rate of cell death of Lewy inclusion-bearing neurons in our in vivo multiphoton imaging paradigm, but PLK2 deletion does slow the rate of neuronal death. Our data indicate that inhibition of PLK2 represents a promising avenue for developing new therapeutics, but that the mechanism of neuroprotection by PLK2 inhibition is not likely due to reducing alpha-synuclein serine-129 phosphorylation and that the true Lewy body kinase still awaits discovery.

Neurovascular coupling is a crucial mechanism that matches the high energy demand of the brain with a supply of energy substrates from the blood. Signaling within the neurovascular unit is responsible for activity-dependent changes in cerebral blood flow. The strength and reliability of neurovascular coupling form the basis of non-invasive human neuroimaging techniques, including blood oxygen level dependent (BOLD) functional magnetic resonance imaging. Interestingly, BOLD signals are negative in infants, indicating a mismatch between metabolism and blood flow upon neural activation; this response is the opposite of that observed in healthy adults where activity evokes a large oversupply of blood flow. Negative neurovascular coupling has also been observed in rodents at early postnatal stages, further implying that this is a process that matures during development. This rationale is consistent with the morphological maturation of the neurovascular unit, which occurs over a similar time frame. While neurons differentiate before birth, astrocytes differentiate postnatally in rodents and the maturation of their complex morphology during the first few weeks of life links them with synapses and the vasculature. The vascular network is also incomplete in neonates and matures in parallel with astrocytes. Here, we review the timeline of the structural maturation of the neurovascular unit with special emphasis on astrocytes and the vascular tree and what it implies for functional maturation of neurovascular coupling. We also discuss similarities between immature astrocytes during development and reactive astrocytes in disease, which are relevant to neurovascular coupling. Finally, we close by pointing out current gaps in knowledge that must be addressed to fully elucidate the mechanisms underlying neurovascular coupling maturation, with the expectation that this may also clarify astrocyte-dependent mechanisms of cerebrovascular impairment in neurodegenerative conditions in which reduced or negative neurovascular coupling is noted, such as stroke and Alzheimer’s disease.