An Ultraconserved Brain-Specific Enhancer Within ADGRL3 (LPHN3) Underpins Attention-Deficit/Hyperactivity Disorder Susceptibility.
Genetic factors predispose individuals to attention-deficit/hyperactivity disorder (ADHD). Previous studies have reported linkage and association to ADHD of gene variants within ADGRL3. In this study, we functionally analyzed noncoding variants in this gene as likely pathological contributors.



In silico, in vitro, and in vivo approaches were used to identify and characterize evolutionary conserved elements within the ADGRL3 linkage region (~207 Kb). Family-based genetic analyses of 838 individuals (372 affected and 466 unaffected patients) identified ADHD-associated single nucleotide polymorphisms harbored in some of these conserved elements. Luciferase assays and zebrafish green fluorescent protein transgenesis tested conserved elements for transcriptional enhancer activity. Electromobility shift assays were used to verify transcription factor-binding disruption by ADHD risk alleles.



An ultraconserved element was discovered (evolutionary conserved region 47) that functions as a transcriptional enhancer. A three-variant ADHD risk haplotype in evolutionary conserved region 47, formed by rs17226398, rs56038622, and rs2271338, reduced enhancer activity by 40% in neuroblastoma and astrocytoma cells (pBonferroni< .0001). This enhancer also drove green fluorescent protein expression in the zebrafish brain in a tissue-specific manner, sharing aspects of endogenous ADGRL3 expression. The rs2271338 risk allele disrupts binding of YY1 transcription factor, an important factor in the development and function of the central nervous system. Expression quantitative trait loci analysis of postmortem human brain tissues revealed an association between rs2271338 and reduced ADGRL3 expression in the thalamus.



These results uncover the first functional evidence of common noncoding variants with potential implications for the pathology of ADHD.
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland. Electronic address: ariel.martinez@nih.gov.
Ariel F
AF
Martinez
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland.
Yu
Y
Abe
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland.
Sungkook
S
Hong
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland.
Kevin
K
Molyneux
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland.
David
D
Yarnell
Institute of Biology I, University of Freiburg, Freiburg, Germany.
Heiko
H
Löhr
Institute of Biology I, University of Freiburg, Freiburg, Germany.
Wolfgang
W
Driever
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland.
Maria T
MT
Acosta
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland; Genome Biology Department, John Curtin School of Medical Research, The Australian National University, Canberra, Australia.
Mauricio
M
Arcos-Burgos
Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland.
Maximilian
M
Muenke
D000293:Adolescent; D000328:Adult; D001289:Attention Deficit Disorder with Hyperactivity / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D005260:Female; D020022:Genetic Predisposition to Disease; D006801:Humans; D008297:Male; D010375:Pedigree; D020641:Polymorphism, Single Nucleotide; D043562:Receptors, G-Protein-Coupled / Q000235:genetics*; D018000:Receptors, Peptide / Q000235:genetics*; D055815:Young Adult
C513284:ADGRL3 protein, human; D043562:Receptors, G-Protein-Coupled; D018000:Receptors, Peptide
ADGRL3; ADHD; Cis-acting regulatory element; Enhancer; Evolutionary conserved regions; Genetics; LPHN3; Latrophilin; Zebrafish
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