Regulation of α-synuclein expression in cultured cortical neurons.

Alpha-synuclein (SNCA) is a predominantly neuronal protein involved in the control of neurotransmitter release. The levels of SNCA expression are closely linked to the pathogenesis of Parkinson's disease; however, the biochemical pathways and transcriptional elements that control SNCA expression are not well understood. We previously used the model system of neurotrophin-mediated PC12 cell neuronal differentiation to examine these phenomena. Although these studies were informative, they were limited to the use of a cell line; therefore, in the current work, we have turned our attention to cultured primary rat cortical neurons. In these cultures, SNCA expression increased with time in culture, as the neurons mature. Luciferase assays based on transient transfections of fusion constructs encoding components of the transcriptional control region of SNCA identified various promoter areas that have a positive or negative effect on SNCA transcription. Intron 1, previously identified by us as an important regulatory region in the PC12 cell model, cooperates with regions 5' to exon 1 to mediate gene transcription. Using selective pharmacological tools, we find that tyrosine kinase receptors and the phosphatidyl-inositol 3 kinase signaling pathway are involved in mediating these effects. The exogenous application of the neurotrophin brain-derived neurotrophic factor (BDNF) is sufficient on its own to promote the transcriptional activation of SNCA through this pathway, but a neutralizing antibody against BDNF failed to affect SNCA transcription in maturing cultures, suggesting that BDNF is not the main factor involved in maturation-induced SNCA transcription in this model. Further in vivo studies are needed to establish the role of neurotrophin signaling in the control of SNCA transcription.

Functional dissection of the alpha-synuclein promoter: transcriptional regulation by ZSCAN21 and ZNF219.

Alpha-synuclein (SNCA) is an abundant neuronal protein involved in synaptic neurotransmission. SNCA expression levels have been strongly implicated in Parkinson's disease pathogenesis. We have previously demonstrated that in the PC12 cell line elements in intron 1 may mediate SNCA transcriptional regulation in response to neurotrophins. We have now identified transcription factor (TF) binding sites in intron 1 and the 5'-promoter of SNCA. A binding site for the TF zinc finger and SCAN domain containing (ZSCAN)21 in the 5'-region of intron 1 is required for intron 1 transcriptional activity. Small interfering RNA against ZSCAN21 inhibits activation in the luciferase assay and diminishes SNCA protein levels in naïve and neurotrophin-treated PC12 cells and in primary cultured cortical neurons, demonstrating that ZSCAN21 is a novel transcriptional regulator of SNCA in neuronal cells. The 5'-promoter of SNCA has a complex architecture, including multiple binding sites for the TF zinc finger protein (ZNF)219, which functions as both an activator and a repressor. Targeting ZSCAN21 or other TFs controlling SNCA transcriptional activity may provide novel therapeutic avenues not only for Parkinson's disease but also for other synucleopathies.