Eric Sean Vornholt

Background: Excessive alcohol consumption has become a growing public health concern worldwide due to the potential development of alcohol dependence (AD). Prolonged alcohol abuse leads to dysregulation of the mesocorticolimbic pathway (MCL), effectively disrupting executive functioning and the allostatic conditioning of reward response. Methods: We utilized weighted gene co-expressed network analysis (WGCNA) and network preservation using a case/control study design ( n =35) to identify unique and shared biological processes dysregulated in AD in the prefrontal cortex (PFC) and nucleus accumbens (NAc). We used correlation and regression analyses to identify mRNA/miRNA interactions and local expression quantitative trait loci (cis-eQTL) to identify genetic regulatory mechanisms for networks significantly associated with AD. Results: Network analyses revealed 6 and 3 significant mRNA modules from the NAc and PFC, respectively. Network preservation revealed immune response upregulation in both regions, whereas cellular morphogenesis/localization and cilia-based cell projection processes were upregulated only in the NAc. We observed 4 significantly correlated module eigengenes (ME) between the significant mRNA and miRNA modules in PFC, and 6 significant miRNA/mRNA ME correlations in NAc, with the mir-449a/b cluster emerging as a potential regulator for cellular morphogenesis/localization dysregulation in this brain region. Finally, we identified cis-eQTLs (37 mRNA and 9 miRNA in NAc, and 17 mRNA and 16 miRNA in PFC) which potentially mediate alcohol’s effect in a brain region-specific manner. Conclusion: In agreement with previous reports, we observed a generalized upregulation of immune response processes in subjects with AD, that highlights alcohol’s neurotoxic properties, while simultaneously demonstrating distinct molecular changes in subcortical brain regions as a result of chronic alcohol abuse. Such changes further support previous neuroimaging and physiological studies that emphasize the distinct roles PFC and NAc play in the development of addictive behaviors.

Recently, long noncoding RNA (lncRNA) were implicated in the etiology of alcohol dependence (AD). As lncRNA provide another layer of complexity to the transcriptome, assessing their expression in the brain is the first critical step towards understanding lncRNA functions in AD. To that end, we profiled the expression of lncRNA and protein coding genes (PCG) in nucleus accumbens (NAc) from 41 subjects with AD and 41 controls. At false discovery rate (FDR) of 5%, we identified 69 and 309 differentially expressed lncRNA and PCG genes, respectively. Using co-expression network analyses, we identified three lncRNA and five PCG modules significantly correlated with AD at Bonferroni adj. p≤0.05. To better understand lncRNA functions, we integrated the lncRNA and PCG hubs from the significant AD modules; at FDR of 5%, we identified 3 151 positive and 2 255 negative correlations supporting the functional role of lncRNA in the development of AD. Gene enrichment analysis revealed that PCG significantly correlated with lncRNA are, among others, enriched for neuronal and immune related processes. To highlight the mechanisms, by which genetic variants contribute to AD, we integrated lncRNA and PCG hubs with genome-wide SNP data. At FDR≤0.3, we identified 276 expression quantitative trait loci (eQTL), affecting the expression of 20 and 256 lncRNA and PCG hubs, respectively. Our study is the first to profile lncRNA expression in nucleus accumbens in a large postmortem alcohol brain sample and our results may provide novel insights into the regulation of the brain transcriptome across disease.