Allele and genotype frequencies, Hardy-Weinberg distribution and analysis of covariance of ATG7 V471A in different European populations.

<p>Populations with ≥100 examined HD patients are shown; <i>n</i> number of investigated HD patients.</p>a<p>Allele frequency of nucleotide substitution in <i>ATG7</i> is described by V ( = valine) as major allele and A ( = alanine) as rare allele.</p>b<p>Compared to the number of investigated patients in the previous study <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0068951#pone.0068951-Metzger3" target="_blank">[20]</a> we managed it to re-genotype 25 HD patients of the 1st European HD cohort, which failed during the first genotyping.</p>c<p>Analysis of covariance showing the effect of the ATG7 V471A polymorphism on the HD AAO in the respective population.</p>*<p>significant effect, the level of significance was set to P = 0.05 and the p-value was adjusted for multiple testing according to Bonferroni correction.</p

Mean ages at onset and CAG repeat numbers of the expanded huntingtin allele in different European populations.

<p>Populations with ≥100 examined HD patients are shown; <i>n</i> number of investigated HD patients.</p><p>AAO, age at onset, CAGexp, expanded CAG repeat number in huntingtin, SD standard deviation, V major allele valine at amino acid position 471 (V471), A rare allele alanine at amino acid position 471 (A471).</p>a<p>t test: Comparing the mean AAO of the VV and VA genotypes in the different populations, only Italian patients with genotype VV differ significantly from patients with heterozygous VA genotype (P = 0.0348).</p>b<p>t-test: Italian patients show the oldest mean AAO, which is significantly older than the mean AAO of patients from the other presented European countries together (Italy vs other countries: P<0.0001), but not significantly different from the mean AAO of patients from Denmark (P = 0.4728) and France (P = 0.2050).</p

Analysis of covariance of ATG7 in EHDN REGISTRY cohort.

<p>The level of significance was set to P = 0.05; n = 1464.</p><p>CAGexp, expanded CAG allele in huntingtin.</p>a<p>Minimum number of patients, which are necessary to detect a significant effect of the analysed factor based on the respective genotypes.</p

RT-qPCR analysis of <i>E2F2</i> gene expression in HD patients, according to <i>E2F2</i> rs2742976 genotype.

<p>Two methods were used. In Taqman assay, the expression of <i>E2F2</i> gene was analyzed in 31 samples (N _TT = 4; N _GT = 12, N _GG = 15) with Hs00918089_m1 Taqman probe; the expression values were normalized respect to expression of <i>B2M</i> and <i>YWHAZ</i> reference genes. In SYBR Green assay, the <i>E2F2</i> gene expression was estimated in 31 samples (N_TT = 5; N_GT = 14, N_GG = 12); the expression values were normalized to expression of <i>UBC</i> and <i>YWHAZ</i> reference genes. Results are expressed as fold over respective GG individuals. Asterisk denotes statistically significant differences (P<0.05) between GG and any other group, according to DataAssist software analysis (T-test) or REST software analysis (Pair Wise Reallocation Randomization test).</p

CAGexp and mAO comparisons between samples from Southern and Northern European populations.

<p>CAGexp and mAO comparisons between samples from Southern and Northern European populations.</p

CAGexp and eAO comparisons between samples from Southern and Northern European populations.

<p>CAGexp and eAO comparisons between samples from Southern and Northern European populations.</p