An integrative approach prioritizes the orphan GPR61 genomic region in tissue-specific regulation of chronotype

Abstract

Study objectives: Chronotype, a manifestation of circadian rhythms, affects morning or evening preferences and ease of getting up. This study explores the genetic basis of morning chronotype and ease of getting up, focusing on the G-protein-coupled receptor locus, GPR61.

Methods: We analyzed the genetic correlation between chronotype and ease of getting up using linkage disequilibrium score regression with summary statistics from the UK Biobank (n = 453,379). We prioritized shared signals between chronotype and ease of getting up using the Human Genetic Evidence (HuGE) score. We assessed the significance of GPR61 and the lead variant rs12044778 through co-localization and in silico analyses from ENCODE, Genotype-Tissue Expression, Hi-C, and Knockout Mouse Project databases to explore potential regulatory roles of causal genes.

Results: We identified a strong genetic correlation (Rg = 0.80, p = 4.9 × 10³²⁴) between chronotype and ease of getting up. Twenty-three genes, including three circadian core clock components, had high HuGE scores for both traits. Lead variant rs12044778 in GPR61 was prioritized for its high HuGE score (45) and causal pleiotropy (posterior probability = 0.98). This morningness variant influenced gene expression in key tissues: decreasing GPR61 in tibial nerve, increasing AMIGO1 in subcutaneous adipose, and increasing ATXN7L2 in the cerebellum. Functional knockout models showed GPR61 knockout increased fat mass and activity, AMIGO1 knockout increased activity, and ATXN7L2 knockout reduced body weight without affecting activity.

Conclusions: Our findings reveal pleiotropic genetic factors influencing chronotype and ease of getting up, emphasizing GPR61's rs12044778 and nearby genes like AMIGO1 and ATXN7L2. These insights advance our understanding of circadian preferences and suggest potential therapeutic interventions.

Keywords: AMIGO1; ATXN7L2; GPR61; circadian rhythm; ease of getting up; genetic pleiotropy; knockout models; morning chronotype; rs12044778.

The genetic regulation of morningness chronotype and wakefulness highlights the role of GPR61 rs12044778 and its systemic effects on adiposity and activity. The model illustrates the genetic and physiological pathways linking the GPR61 gene variant rs12044778 to chronotype preferences and the ease of waking up. It depicts the variant’s regulatory effects on GPR61 expression in the tibial nerve and its downstream effects on other genes like AMIGO1 and ATXN7L2 in brain regions and adipose tissue. The consequences of gene knockouts in mouse models are shown, with alterations in adiposity and activity levels, mapping the genetic influence to observable phenotypes.

Figure 1.

GPR61 is associated with chronotype and ease of waking up. The plot displays the Human Genetic Evidence (HuGE) score calculated to quantify the genetic support of the involvement of GPR61 (box) in GWAS traits (SleepChronotype: Morningness Chronotype and EaseOfWakingUp: Ease of Waking up in the morning). The canonical clock genes PER3, PER2, and CRY1 are highlighted by a brown box. The score was calculated from GWAS significant loci with a p-value cutoff of 5e−8.

Figure 2.

Co-localization analysis reveals a shared variant at the GPR61 locus in GWAS of morningness chronotype and ease of waking up. Regional association plot at the colocalized GPR61 locus with GWAS significance (p-value < 5e−8) for (A) morningness chronotype and (B) ease of waking up. The y-axis shows the −log10 p-value for each variant in the region, and the x-axis shows the genomic position. Each variant is represented by a filled circle, with the rs12044778 lead variant as a diamond and nearby variants colored according to the degree of linkage disequilibrium (r²) with rs12044778 in the 1000G EUR population. The lower panel shows genes located in the displayed region, and the blue line corresponds to the recombination rate. The Human Genetic Evidence (HuGE) score (C) for the genes in the genomic region of the lead variant rs12044778 Chr1: 109,832,494–110,341,028 (Genome Build: hg19/GRCh37).

Figure 3.

Association of the co-localized lead variant rs12044778 with accelerometer-derived sleep and activity timing measures. The plot displays the variant-level phenotypic associations for the colocalized lead variant rs12044778 expressed as effect size (Beta; y-axis) and significance (–log10(p-value)). The dashed line is the significance threshold of 0.005 (0.05/10 Circadian Phenotypes). Morning Chrono: Morningness Chronotype; Ease of Getup: Ease of waking up in the morning; M10: Most-active 10-hour timing; L5: least-active 5-hour timing, and M5: Sleep Midpoint timing, SE: Sleep efficiency, MSD: Mean sleep duration, SDSD: Standard deviation o of sleep duration, NSE: Number of sleep episodes, DID: Diurnal inactivity duration (Table S1).

Figure 4.

The lead variant rs12044778 in GPR61 regulates the gene expression of nearby genes. Plot (A) displays the transcription factors whose binding is predicted to be affected by the variant rs12044778, and it is expressed in delta, which is the binding affinity between the reference allele and the alternate allele for a specific TF binding motif. (B) shows rs12044778 is at the binding site of EGR-1, ELF-1, GABPA, and NR2F2 transcription factors using K562 cell Chip-Seq experiment from ENCODE. Plot (C) is displayed as normalized gene expression and rs12044778 genotype in Gtex v.8 data with (the number of participants) for each genotype below it. The variant rs12044778 decreases the gene expression of GPR61 in nerve tibial, increases AMIGO1 expression in subcutaneous adipose tissue, and increases ATXN7L2 gene expression in the cerebellum.

Figure 5.

The tissue-specific interaction of the lead variant rs12044778 with nearby genes. Hi-C Interaction analysis of the rs12044778 locus in Adipose-Progenitor Mesenchymal Stem Cells (A) and astrocyte of the cerebellum (B). (A) Heatmap depicting the Hi-C contact map of chromosome 1, highlighting the interaction frequencies within a specific genomic region. Topologically associating domains (TADs) are indicated by triangles, demarcating the structural units of chromosomal folding. (B) Bar and dot plots represent the bias-removed interaction frequencies (bars) and their corresponding distance-normalized interaction frequencies (dots), providing a detailed view of the chromatin interactions within and across the TAD boundaries. (C) Chromosome ideogram with gene annotations (D) and blue interaction arcs. The arcs visualize physical contacts between regions, elucidating the three-dimensional organization of the genome in relation to gene regulatory elements and the SNP rs12044778.

Figure 6.

The expression profile of nearby genes regulated by rs12044778 in humans. (A) The bar graph is the gene expression in nTPM of GPR61, GNAI3, AMIGO1, and ATXN7L2 gene expression in the subcutaneous adipose tissues (ADIP) hippocampus (HIP), pituitary gland (PIT), cerebellum (CER), and hypothalamus (HYP). (B) The box-whisker plot is the gene expression in nTPM of GPR61, GNAI3, AMIGO1, and ATXN7L2 gene expression the hypothalamic the arcuate nucleus (ARC), dorsomedial hypothalamic nucleus (DMH), lateral hypothalamic area (LHA), ventromedial hypothalamic nucleus (VMH), and preoptic area (POA).