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. Author manuscript; available in PMC: 2016 Oct 20.
Published in final edited form as: Nature. 2016 Apr 20;533(7601):95–99. doi: 10.1038/nature17939

Fig. 3. Sequence-specific effect of PD-associated risk variants on binding of CNS expressed TFs EMX1 and NKX6-1 at SNCA intron-4 enhancer.

Fig. 3

(a) ChIP-qRT-PCR analysis for binding of indicated TFs at the intron-4-enhancer element compared with a negative control region in the SNCA locus (calculated as fold enrichment compared with IgG Isotype control). Statistical significance was determined using t-test followed by the Holm-Šídák method to correct for multiple comparisons (alpha = 0.05). *P < 0.0001. (b, c) EMSA analysis for SNP-genotype-specific binding of (b) EMX2 or (c) NKX6-1 to oligonucleotides (oligo) harboring the indicated genotype at rs356168 (A/G-allele). Binding was analyzed in nuclear extracts (NEs) from wild-type (293) or EMX1 (b, EMX) or NKX6-1 (c, NKX) overexpressing HEK293 cells. Red arrows point to oligonucleotide-specific binding which is lost in the presence of unlabeled competitor oligonucleotides (indicated genotype at rs356168; 200X). (d) SNCA expression analysis following doxycycline (DOX)-induced overexpression of EGFP, EMX2 or NKX6-1 for 3 days in terminally differentiated neurons (differentiation day 21). Shown are mean values ± s.d. (n = 10) of relative SNCA expression in DOX-induced cells compared with the corresponding untreated controls (NoDOX). Results are representative of two different experiments. Statistical significance was calculated using t-test followed by the Holm-Šídák method to correct for multiple comparisons (alpha = 0.05). *P < 0.0001. Source data are provided as Source Data for Extended Data Figure 3.