Top 30 genes significantly more highly expressed in the RPE compared to the IE.

<p>The genes that are in bold were shown to be enriched in the human RPE [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0182983#pone.0182983.ref026" target="_blank">26</a>]. Asterisks mark the genes that might be present in our dataset by contamination of the mRNA on the photoreceptor-RPE interface or may be expressed to some extent in both adjacent cell layers (also see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0182983#sec021" target="_blank">Materials and methods</a>).</p

Top 20 significant canonical pathways of the core analysis in IPA (Ingenuity) of most highly expressed genes of the IE and the RPE.

<p>P-values indicate the significance of enrichment for the most highly expressed genes from our dataset. P-values were corrected for multiple testing using the Benjamini-Hochberg (B-H) false discovery rate. The upper graph (light blue bars) represents the–log(B-H) p-value of the RPE and the lower graph (dark blue bars) represents the–log(B-H) p-value of the IE. The orange line indicates the threshold of B-H corrected p<0.001.</p

Heatmap for the expression of genes related to RPE specific functions.

<p>The normalized expression data are converted to heat map color using the mean and maximum values for each gene. The intensity scale of the standardized expression values ranges from dark blue (low expression) to dark orange (high expression). We added a hierarchical cluster tree that shows that the IE samples cluster together and the RPE samples cluster together.</p

Post-mortem brain material used in this study.

<p>Listed are clinical diagnosis (CON = control, AD = Alzheimer’s disease), Braak stage, gender, age (in years), ApoE genotype, post-mortem interval (PMI, in hours: minutes).</p

QC expression is not upregulated by Aβ or the UPR.

<p>A. Differentiated SK-N-SH cells were treated with 1 µM and 2 µM oligomeric (O) and fibrillary (F) Aβ_1–42 for 24 h. Shown is the average + SD of normalized QC mRNA levels of triplicates from a representative experiment B. Differentiated SK-N-SH cells were treated with different UPR inducers, TM (0.2 µg/µl and 0.5 µg/µl), TG (1 µM and 2 µM) and DD (20 mM) for 16 h. Shown is the average + SD of normalized QC mRNA levels of n = 9 from 3 independent experiments. Normalized BiP mRNA levels are shown as positive control for UPR induction (n = 6 from two independent experiments). The expression levels were normalized to eEF2α mRNA, the levels in untreated cells are set to 1. Asterisks indicate a significant difference compared with control (*p≤0.01, **p≤0.001, ***p≤0.0001).</p

Disturbed Ca²⁺ homeostasis increases QC expression and enzyme activity.

<p>A. Differentiated SK-N-SH cells were treated with different concentrations TG as indicated. Shown are the average + SD of normalized QC mRNA levels of n = 6 from two independent experiments. The expression levels were normalized to eEF2α mRNA, the expression levels in untreated cells are set to 1. B. Differentiated SK-N-SH cells were treated with 1 µM TG for 16 h. The enzymatic activity of QC activity was determined in protein lysates as described in materials and methods, activity in untreated cells is set to 1. Shown is average +SEM of n = 15 from 5 independent experiments. Asterisks indicate a significant difference compared to control (*p≤0.001, **p≤0.0001).</p

Induction of c-fos and c-jun precedes increased QC expression.

<p>Differentiated SK-N-SH cells were treated with 1 µM TG for the times indicated. Shown is the average + SD of normalized QC (A), c-fos (B) and c-jun (C) mRNA levels of triplicates from a representative experiment of three. The expression levels were normalized to eEF2α mRNA, the expression levels in untreated cells are set to 1. Asterisks indicate a significant difference compared to control (*p<0.05; **p≤0.001; ***p≤0.0001).</p

Primers and probes used for qPCR 480 light cycler.

<p>Probe numbers refer to numbers in the Roche universal probe library.</p

ER Ca²⁺ depletion increases QC levels.

<p>Differentiated SK-N-SH cells were treated with 1 µM TG for 16 h with or without pre-incubation with 5 µM BAPTA-AM (B) for 1 h. Shown are the average + SD of normalized QC mRNA levels of n = 9 from 3 independent experiments. The expression levels were normalized to eEF2α mRNA, the expression levels in untreated cells are set to 1. Asterisks indicate a significant difference compared to control (*p≤0.0001).</p

QC mRNA expression is increased in earliest stages of AD pathology.

<p>Expression of QC mRNA was determined in a cohort of patients with varying stages of AD pathology (see materials and methods for details). CON and AD refer to the clinical diagnosis, BS is Braak score for tau pathology, PL is the plaqueload in the hippocampus/entorhinal cortex. Shown is a box-plot of results of the pathological groups as indicated in hippocampus/entorhinal cortex. The expression levels were normalized to eEF2α mRNA. Kruskall-Wallis test was used to evaluate differences between groups followed by the Mann-Whitney U test, to test differences between pairs of groups.</p