RT Journal Article SR Electronic T1 CRISPR disruption and UK Biobank analysis of a highly conserved polymorphic enhancer suggests a role in male anxiety and ethanol intake JF bioRxiv FD Cold Spring Harbor Laboratory SP 572065 DO 10.1101/572065 A1 McEwan, Andrew R. A1 Davidson, Connor A1 Hay, Elizabeth A1 Turnbull, Yvonne A1 Erickson, Johanna Celene A1 Marini, Pietro A1 Wilson, Dana A1 McIntosh, Andrew M. A1 Adams, Mark J. A1 Murgatroyd, Chris A1 Barrett, Perry A1 Delibegovic, Mirela A1 Clarke, Toni-Kim A1 MacKenzie, Alasdair YR 2019 UL http://biorxiv.org/content/early/2019/12/13/572065.abstract AB Excessive alcohol intake is associated with 5.9% of global deaths. However, this figure is especially acute in men such that 7.6% of deaths can be attributed to alcohol intake. Previous studies identified a significant interaction between genotypes of the galanin (GAL) gene with anxiety and alcohol abuse in different male populations but were unable to define a mechanism. To address these issues the current study analysed the human UK Biobank cohort and identified a significant interaction (n=115,865; p=0.0007) between allelic variation (GG or CA genotypes) in the highly conserved human GAL5.1 enhancer, alcohol intake (AUDIT questionnaire scores) and anxiety in men that was consistent with these previous studies. Critically, disruption of GAL5.1 in mice using CRISPR genome editing significantly reduced GAL expression in the amygdala and hypothalamus whilst producing a corresponding reduction in ethanol intake in KO mice. Intriguingly, we also found evidence of reduced anxiety-like behaviour in male GAL5.1KO animals mirroring that seen in humans. Using bioinformatic analysis and co-transfection studies we further identified the EGR1 transcription factor, that is co expressed with GAL in amygdala and hypothalamus, as being important in the protein kinase C (PKC) supported activity of the GG genotype of GAL5.1 but less so in the CA genotype. Our unique study uses a novel combination of human association analysis, CRISPR genome editing in mice, animal behavioural analysis and cell culture studies to identify a highly conserved regulatory mechanism linking anxiety and alcohol intake that might contribute to increased susceptibility to anxiety and alcohol abuse in men.