Summary:
α-synucleinis a small presynaptic neuronal protein, encoded by the SNCA gene,
that is implicated genetically and neuropathologically in Parkinson’s disease
(PD). A large body of evidence has established that PD pathogenesis is closely
linked to increased levels of SNCA; however to date, the biochemical pathways
and transcriptional elements that control SNCA expression are still obscure.
Previous experiments in our laboratory in the PC12 cell line demonstrated that
the transcription factor ZSCAN21 binds to the intron 1 region of the Snca gene
and is strongly involved in its transcriptional regulation. Therefore, in the
current experiments, we wished to characterize further the role of ZSCAN21 in
Snca transcriptional regulation in primary cultures and in vivo. We find that
in vivo ZSCAN21 is expressed in neurons and its levels are developmentally
regulated in different brain regions where ASYN is also detected. Further, we
confirmed through Chromatin Immunoprecipitation its presence in a binding
complex in the intron 1 region of the Snca gene in rat cortical neuronal
cultures. Importantly, lentiviral-mediated silencing of Zscan21 increased
significantly the promoter activity of Snca as well as its mRNA and protein
levels in such cultures. In contrast, Zscan21mediated silencing in
differentiated neurosphere cultures reduced Snca levels. Stereotaxic delivery
of adeno-associated virus against Zscan21in the postnatal and adult
hippocampus, an area linked with the non-motor symptoms of PD, revealed no
significant alterations in Snca levels, despite efficient Zscan21 knock-down.
Interestingly, Zscan21overexpression in cortical neurons with adenoviruses led
to robust mRNA but negligible protein expression, suggesting that ZSCAN21
protein levels are tightly regulated post-transcriptionally and / or
post-translationally. Therefore, overall, our study demonstrates that ZSCAN21,
a transcription factor whose levels are under strict posttranscriptional /
posttranslational control in neurons, isdiversely implicated in the
transcriptional regulation of Snca in respect to the developmental stage, at
least in in vitro primary neuronal settings. In vivo, however, the unaltered
Snca levels observed following Zscan21downregulation, imply the presence of
alternative or perhaps compensatory mechanisms that regulate Snca transcription
in such settings. Furthermore, in a genetic case control study of the ZSCAN21
binding site in SNCA intron1, we did not find polymorphisms between PD patients
and controls, suggesting that genetic diversity within this region does not
contribute to disease pathogenesis. Overall, given the diverse effects in cell
culture, and the lack of discernible in vivo effects, and although further
studies are needed, our work does not provide sufficient support for the idea
of targeting ZSCAN21 in order to manipulate SNCA levels in synucleinopathy
models.
Keywords:
Parkinson's disease, α-synuclein, ZSCAN21 transcription factor, Transciptional regulation, Rat
