Integrating mRNA and miRNA Weighted Gene Co-Expression Networks with eQTLs in the Nucleus Accumbens of Subjects with Alcohol Dependence.

Alcohol consumption is known to lead to gene expression changes in the brain. After performing weighted gene co-expression network analyses (WGCNA) on genome-wide mRNA and microRNA (miRNA) expression in Nucleus Accumbens (NAc) of subjects with alcohol dependence (AD; N = 18) and of matched controls (N = 18), six mRNA and three miRNA modules significantly correlated with AD were identified (Bonferoni-adj. p≤ 0.05). Cell-type-specific transcriptome analyses revealed two of the mRNA modules to be enriched for neuronal specific marker genes and downregulated in AD, whereas the remaining four mRNA modules were enriched for astrocyte and microglial specific marker genes and upregulated in AD. Gene set enrichment analysis demonstrated that neuronal specific modules were enriched for genes involved in oxidative phosphorylation, mitochondrial dysfunction and MAPK signaling. Glial-specific modules were predominantly enriched for genes involved in processes related to immune functions, i.e. cytokine signaling (all adj. p≤ 0.05). In mRNA and miRNA modules, 461 and 25 candidate hub genes were identified, respectively. In contrast to the expected biological functions of miRNAs, correlation analyses between mRNA and miRNA hub genes revealed a higher number of positive than negative correlations (χ2 test p≤ 0.0001). Integration of hub gene expression with genome-wide genotypic data resulted in 591 mRNA cis-eQTLs and 62 miRNA cis-eQTLs. mRNA cis-eQTLs were significantly enriched for AD diagnosis and AD symptom counts (adj. p = 0.014 and p = 0.024, respectively) in AD GWAS signals in a large, independent genetic sample from the Collaborative Study on Genetics of Alcohol (COGA). In conclusion, our study identified putative gene network hubs coordinating mRNA and miRNA co-expression changes in the NAc of AD subjects, and our genetic (cis-eQTL) analysis provides novel insights into the etiological mechanisms of AD

(A). rs11626307 effect on the hsa-miR-134-5p (A) and hsa-miR-370-3p (B) expressions.

<p>(A). rs11626307 effect on the hsa-miR-134-5p (A) and hsa-miR-370-3p (B) expressions.</p

eQTL (chr2:170783092:D) effect on the expression of the glutamate decarboxylase (GAD1) gene.

<p>The bar plot depicts the differential expression of GAD1 among homozygote for the major (11, red), heterozygote (12, green) and homozygote for the minor alleles (22, blue) subjects.</p

(A) Cluster dendrogram and module assignment for mRNA modules from WGCNA. Topological overlap dissimilarity measure is clustered by average linkage hierarchical clustering and module assignments (dynamic hybrid algorithm) are denoted in the color bar (bottom). 4571 transcripts were assigned to one of 24 modules including Mgrey. (B). Following the same outline, 240 miRNAs are assigned to one of 12 modules indicated by color (including Mgrey).

<p>(A) Cluster dendrogram and module assignment for mRNA modules from WGCNA. Topological overlap dissimilarity measure is clustered by average linkage hierarchical clustering and module assignments (dynamic hybrid algorithm) are denoted in the color bar (bottom). 4571 transcripts were assigned to one of 24 modules including Mgrey. (B). Following the same outline, 240 miRNAs are assigned to one of 12 modules indicated by color (including Mgrey).</p

Module-trait relationships.

<p>(<b>A</b>) mRNA module MEs are correlated (Pearson) to AD case-status (Class), brain pH, PMI, Age, RIN and subject smoking status to assess for confounding. P-values shown are unadjusted for multiple testing. After adjusting for number of modules tested, ME_{<i>turquoise</i>}, ME_{<i>yellow</i>}, ME_{<i>grey60</i>}, ME_<i>pink</i>, ME_<i>green</i> and ME_{<i>salmon</i>} are significantly correlated with AD case-status (Class). (<b>B</b>). Similarly, after adjusting p-values for number of modules tested, miRNAs ME_{<i>yellow</i>}, ME_<i>blue</i> and ME_<i>brown</i> modules are significantly correlated with AD case-status (Class).</p

Brain list enrichment for cell type specific modules.

<p>Brain list enrichment for cell type specific modules.</p