Multiomic analyses direct hypotheses for Creutzfeldt-Jakob disease risk genes

Abstract

Prions are assemblies of misfolded prion protein that cause several fatal and transmissible neurodegenerative diseases, with the most common phenotype in humans being sporadic Creutzfeldt-Jakob disease (sCJD). Aside from variation of the prion protein itself, molecular risk factors are not well understood. Prion and prion-like mechanisms are thought to underpin common neurodegenerative disorders meaning that the elucidation of mechanisms could have broad relevance. Herein we sought to further develop our understanding of the factors that confer risk of sCJD using a systematic gene prioritization and functional interpretation pipeline based on multiomic integrative analyses. We integrated the published sCJD genome-wide association study summary statistics with publicly available bulk brain and brain cell type gene and protein expression datasets. We performed multiple transcriptome and proteome-wide association studies and Bayesian genetic colocalization analyses between sCJD risk association signals and multiple brain molecular quantitative trait loci signals. We then applied our systematic gene prioritization pipeline to the obtained results and nominated prioritized sCJD risk genes with risk-associated molecular mechanisms in a transcriptome and proteome-wide manner. Genetic upregulation of both gene and protein expression of syntaxin-6 (STX6) in the brain was associated with sCJD risk in multiple datasets, with a risk-associated gene expression regulation specific to oligodendrocytes. Similarly, increased gene and protein expression of protein disulfide isomerase family A member 4 (PDIA4), involved in the unfolded protein response, was linked to increased disease risk, particularly in excitatory neurons. Protein expression of mesencephalic astrocyte derived neurotrophic factor (MANF), involved in protection against endoplasmic reticulum stress and sulfatide binding (linking to the enzyme in the final step of sulfatide synthesis, encoded by sCJD risk gene GAL3ST1), was identified as protective against sCJD. In total 32 genes were prioritized into two tiers based on the level of evidence and confidence for further studies. This study provides insights into the genetically-associated molecular mechanisms underlying sCJD susceptibility and prioritizes several specific hypotheses for exploration beyond the prion protein itself, as well as beyond the previously highlighted sCJD risk loci, through the newly prioritized sCJD risk genes and mechanisms. These findings highlight the importance of glial cells, sulfatides and the excitatory neuron unfolded protein response in sCJD pathogenesis.

Keywords: multiomics; neurodegeneration; proteome-wide association studies (PWAS); sporadic Creutzfeldt-Jakob disease (sCJD); transcriptome-wide association studies (TWAS).

Figure 1

Gene prioritization results for sCJD GWAS. A visual summary of weighted evidence category scores for each prioritized risk gene, together with brain cell-type-specific gene expression proportions. The figure shows a total of 32 prioritized risk genes (17 tier 1 and 15 tier 2). The leftmost squares indicate the locus indexes where ‘G’ is used for the genome-wide significant loci, ‘S’ for the subthreshold loci and ‘O’ for the other remaining loci. The types of evidence for each category are coloured according to the three different domains to which they belonged. Weighted scores for each evidence category are rescaled to a 0–100 scale based on the maximum score a candidate gene can obtain from a category (see Supplementary Table 3). The darker colours represent higher scores in categories or higher average gene expression proportions in the seven major brain cell types, while tier 1 prioritized genes are displayed in dark green and tier 2 prioritized genes are displayed in light green. Only tier 1 and tier 2 genes are shown for each locus, and all candidate genes considered and scored can be found in Supplementary Table 4. CADD (v1.7) PHRED scores for index variants are labelled in white within the respective squares in variant annotation domain. coloc = colocalization; COPs = committed oligodendrocyte precursors; ct-eQTL = cell-type-specific eQTL; eQTL = expression QTL; OPCs = oligodendrocyte precursor cells; pQTL = protein-expression QTL; PWAS = proteome-wide association study; SEA-AD = Seattle Alzheimer’s Disease Cell Atlas; TWAS = transcriptome-wide association study.

Figure 2

Regulation of STX6 brain gene and protein expression by the sCJD-risk-colocalized eQTLs and pQTLs within the STX6 locus. The regional plots of (i) sCJD GWAS association signal (n = 17 679), (ii) STX6 brain pQTL signal in DLPFC (ROSMAP DLPFC pQTL catalogue, n = 376), (iii) STX6 bulk brain eQTL signal in DLPFC (ROSMAP DLPFC eQTL catalogue, n = 560), and (iv) STX6 ct-eQTL signal in oligodendrocytes (Bryois et al. ct-eQTL catalogue, n = 192) are shown for 100 kb extended genomic coordinates of the STX6 locus index variant rs3747957 (chr1:180884717-181084717). Boxes in each panel show QTL-GWAS coloc PP4 values between the molecular QTL signal and the GWAS signal for all tested variants (see Supplementary Tables 5 and 6). The index variant is shown in purple, and LD r² values [calculated within 1000 Genome Project non-Finnish European samples (n = 404) with respect to the index variant] are indicated on a colour scale. Variants that are not available in the LD reference panel are shown in grey. y-axis, −log10 GWAS or QTL P; x-axis, GRCh38 genomic position on chromosome 1, together with the annotation for the genomic positions of the protein-coding genes in the locus. coloc = colocalization; ct-eQTL = cell-type-specific eQTL; DLPFC = dorsolateral prefrontal cortex; eQTL = expression QTL; GWAS = genome-wise association panel; LD = linkage disequilibrium; pQTL = protein-expression QTL; QTL = quantitative trait loci; sCJD = sporadic Creutzfeldt-Jakob disease.

Figure 3

sCJD brain proteome-wide association study results. sCJD brain proteome-wide association study (PWAS) results are shown proteome-wide for both of the PWAS reference panels with two mirrored Manhattan plots on the x-axis; the upper side of the plot displays the results for ROSMAP DLPFC PWAS, while the lower side of the plot displays the results for Banner DLPFC PWAS. Proteome-wide significance thresholds (0.05 divided by number of tested associations; see Supplementary Table 2) for both analyses are indicated with red dashed lines and suggestive significance thresholds (1 divided by number of tested associations) with a blue dotted line. All the genes whose protein products pass these thresholds are labelled and coloured based on their significance (red: proteome-wide significant, blue: suggestive significant). The directionality of Z-scores of each PWAS association is represented with upward-pointing triangles (positive Z-score) and downward-pointing triangles (negative Z-score). y-axis, −log10 PWAS P; x-axis, GRCh38 chromosomal positions. DLPFC = dorsolateral prefrontal cortex; sCJD = sporadic Creutzfeldt-Jakob disease.

Figure 4

Speculative model of the cell types and the potential relationship between prioritized risk genes and their mechanisms. A common amino acid variant (p.Val29Met) in the GAL3ST1 gene, encoding galactosylceramide sulfotransferase, increases sulfatide production predominantly in oligodendrocytes, conferring increased risk of sporadic Creutzfeldt-Jakob disease (1). Sulfatide may act as a cofactor in PrP^C conversion or prion propagation, which may be intercepted by the astrocyte-secreted factor, MANF, which binds to sulfatide extracellularly (2). Sulfatide may additionally promote the cellular uptake of MANF allowing it to work in concert with PDIA4 to protect against the adverse effects of endoplasmic reticulum stress and the sustained unfolded protein response characteristic of prion infection (3). Increased syntaxin-6 expression predominantly in oligodendrocytes may be altering the trafficking of either PrP^C or prions with implications on prion formation, propagation and/or clearance (4). Created in BioRender. One, S. (2025) https://BioRender.com/z36z395