Genome-wide methylation in alcohol use disorder subjects: implications for an epigenetic regulation of the cortico-limbic glucocorticoid receptors (NR3C1)

Abstract

Environmental factors, including substance abuse and stress, cause long-lasting changes in the regulation of gene expression in the brain via epigenetic mechanisms, such as DNA methylation. We examined genome-wide DNA methylation patterns in the prefrontal cortex (PFC, BA10) of 25 pairs of control and individuals with alcohol use disorder (AUD), using the Infinium® MethylationEPICBeadChip. We identified 5,254 differentially methylated CpGs (pnominal<0.005). Bioinformatic analyses highlighted biological processes containing genes related to stress adaptation, including the glucocorticoid receptor (encoded by NR3C1). Considering that alcohol is a stressor, we focused our attention on differentially methylated regions of the NR3C1 gene and validated the differential methylation of several genes in the NR3C1 network. Chronic alcohol drinking results in a significant increased methylation of the NR3C1 exon variant 1H, with a particular increase in the levels of 5-hydroxymethylcytosine over 5-methylcytosine. These changes in DNA methylation were associated with reduced NR3C1 mRNA and protein expression levels in PFC as well as other cortico-limbic regions of AUD subjects when compared to controls. Furthermore, we show that the expression of several stress-responsive genes (e.g., CRF, POMC, FKBP5) is altered in the PFC of AUD subjects. These stress-response genes were also changed in the hippocampus, a region that is highly susceptible to stress. These data suggest that alcohol-dependent aberrant DNA methylation of NR3C1 and consequent changes in other stress-related genes might be fundamental in the pathophysiology of AUD and lay the groundwork for treatments targeting the epigenetic mechanisms regulating NR3C1 in AUD.

DNA methylation signature on phosphatidylethanol, not on self-reported alcohol consumption, predicts hazardous alcohol consumption in two distinct populations

“…For example, cg18362496 in H19 that was previously reported hypermethylation in AUD brain samples [61] showed a positive association with PEth in our blood sample (p = 0.03). A stress-related gene, KCNK6, that was previously associated with AUD in brain [31] was nominal significant in the same direction in our study (p = 0.04). However, the majority of significant CpGs for alcohol consumption differs between brain and blood samples.…”

“…Epigenome-wide association studies (EWAS) have identified hundreds of DNAm CpGs in blood for alcohol consumption [25][26][27][28], alcohol use disorder (AUD) [29,30], stress-related alcohol consumption [31], and fetal alcohol syndrome [32][33][34][35]. A large number of CpGs in the blood have recently been reported to have associations with dietary folate and alcohol intake [36].…”

“…Because PCSK9 is well known to regulate low-density lipoprotein cholesterol, DNAm alteration and dysfunction of PCSK9 is thought to be a mechanism for alcoholrelated abnormalities in lipid metabolism. Most recently, Gatta et al [31] reported the hypermethylation of DNA 5methylcytosine at the promoter regions of NR3C1 (Nuclear Receptor Subfamily 3 Group C Member 1), the glucocorticoid receptor, that was correlated with the reduction of mRNA expression of NR3C1 in human brains with AUD. The expression levels of several stressresponsive genes within the NR3C1 gene network were also decreased in brain samples from individuals with AUD.…”

Epigenetic Regulation of GABAergic Neurotransmission and Neurosteroid Biosynthesis in Alcohol Use Disorder

“…Mounting studies have shown evidence for a role for epigenetic mechanisms (e.g., DNA hypermethylation) in the pathophysiology of AUD and associated maladaptive behaviors ( Warnault et al, 2013 ; Berkel and Pandey, 2017 ; Gatta et al, 2017 , 2019 ). In a previous study in the cerebellum of AUD patients, we reported a marked decrease in the amounts of the GABA A R δ subunit, which was associated with increased DNA methylation of the corresponding gene promoter ( Gatta et al, 2017 ).…”

Monoamine and genome-wide DNA methylation investigation in behavioral addiction

“…Indeed, similar processes are thought to take place in BA, which is essentially initiated by environmental stimuli in adulthood or during development. Recent studies with genome-wide DNA methylation assays have unveiled alterations in methylation patterns in drug addiction patients compared with healthy subjects 28 , 29 . DNA methylation is an epigenetic process that regulates gene expression, and as such, it was thought that CpG sites on promoter regions play crucial roles in inhibiting gene expression 30 .…”