Genome- and transcriptome-wide splicing associations with alcohol use disorder.

paper Cited Public

Authors: Huggett, Spencer B; Ikeda, Ami S; Yuan, Qingyue; Benca-Bachman, Chelsie E; Palmer, Rohan H C
Year: 2023
Journal: Scientific reports
PMID: 36894673
DOI: 10.1038/s41598-023-30926-z
PMCID: PMC9998611

Figure 1

Schematic representation of our study. Created with BioRender.com.

Figure 2

Genetic risk for AUD and alternative mRNA Splicing. (A) Violin plot showing polygenic score distributions of AUD between individuals with AUD and controls. Mean and standard error are shown. P-value thresholds were 0.0183, 0.1016, 0.3522 and 0.0187 for the BLA, CEA, NAc and PFC, respectively. Scores were combined across brain regions and tested between AUD cases and controls. (B) Heat matrix showing the amount of variance explained (R2) by polygenic score analyses of differential splicing results for each brain region. Principal components (PC) analysis was used to distil differential splicing results into a single metric (1st PC). Polygenic score p-value thresholds were 5.e−5, 0.006, 5e−5 and 0.0012 for the BLA, CEA, NAc and PFC, respectively.

Figure 3

Alternative mRNA splicing associations with AUD by brain region. (A) Volcano plot displaying differentially spliced genes between individuals with AUD and controls for each brain region. (B) Scatter plot showing differential splicing associations across brain regions from differentially spliced genes. Note ΔPSI stands for the change in percent-spliced-in and that each colored dot represents a specific splicing event in a cluster from a significantly differentially spliced gene (padj < 0.05).

Figure 4

Individual DNA markers linked with alternative mRNA splicing events associated with AUD. (A) Volcano plot showing results from our sQTL analyses. Each dot above the dashed red line represents a significant (padj < 0.05) SNP association with a differentially spliced gene. (B) Bar plot showing the genomic regions enriched for significant sQTL associations. * indicated that a certain genomic region survived correction for multiple testing (padj < 0.05).

Figure 5

SNPs within and around differentially spliced genes contribute to the heritability of AUD. Heritable factors include the observed heritability from LD score regression analyses. Splicing genes include all biallelic SNPs within and 1 Mb around the transcription start and end site of differentially spliced genes associated with AUD in the brain.

#	Section	Preview
0	Introduction	Alternative mRNA splicing is the process where a single gene codes for multiple mRNA transcripts and…
1	Introduction	Alcohol consumption induces abnormal alternative splicing events. That is, in tightly controlled…
2	Introduction	Other research has identified links between genetic predispositions and RNA associations from human…
3	Introduction	Common forms of genetic variation, like single nucleotide polymorphisms (SNPs; individual DNA…
4	Introduction	Our exploratory study hypothesized that (1) polygenic scores of AUD would be higher in those with…
5	Materials and methods — Samples — RNA-seq	We used the same publicly available data source of human post-mortem brain samples as Van Booven et…
6	Materials and methods — Samples — RNA-seq	decreases the portability of polygenic score analyses. Of the human brain samples 23.22% were female…
7	Materials and methods — Samples — RNA-seq	RNA-seq data see Rao et al.14. Briefly, RNA was extracted via the Qiagen RNeasy kit. RNA was…
8	Materials and methods — Samples — RNA-seq	Male primate samples came from four cohorts (4, 5, 7a and 7b) of Rhesus Macaques from the Monkey…
9	Materials and methods — Samples — RNA-seq	Primate samples had five drinking categories: controls (alcohol naïve), low drinkers, high…
10	Materials and methods — Samples — RNA-seq	and Walter et al.16. Briefly, paired-end and stranded RNA libraries were prepared via the TruSeq RNA…
11	Materials and methods — Samples — GWAS	Alternative mRNA splicing associations were inferred from GWAS summary statistics from a study on…
12	Materials and methods — Samples — Data preparation	RNA-seq data were processed using a uniform pipeline. First, we investigated RNA-seq data quality…
13	Materials and methods — Samples — Data preparation	DNA genotypes from human RNA-seq data were ascertained via the SAMtools mpileup function as done…
14	Materials and methods — Samples — Data preparation	resulted in 570,755 SNPs, 178,598 of which passed QC. These ~ 170 k variants were used for polygenic…
15	Materials and methods — Analyses — Differential splicing	To detect alternative mRNA associations with AUD we used Leafcutter version 0.2.923. Leafcutter is a…
16	Materials and methods — Analyses — Differential splicing	intron, < 3 samples per group, and required at least 20 reads, which resulted in 18,685 unique genes…
17	Materials and methods — Analyses — Differential splicing	To assess the overlap between human and primate results we used a Fisher’s Exact test at the…
18	Materials and methods — Analyses — Polygenic scores	We investigated two questions with polygenic score analyses. First, did AUD cases have higher mean…
19	Materials and methods — Analyses — sQTLs	A splicing quantitative trait locus (sQTL) is a SNP that predicts alternative mRNA splicing…

Citation	PMID	DOI	Status
Acosta, G et al., Front. Psychiatry, 2011, Alternative splicing of AMPA subunits in prefrontal cortical fields of cynomolgus monkeys following chronic ethanol self-administration	22291662	10.3389/fpsyt.2011.00072	Cited
Baker, EJ et al., Alcohol. Clin. Exp. Res., 2014, Chronic alcohol self-administration in monkeys shows long-term quantity/frequency categorical stability	25421519	10.1111/acer.12547	Cited
Barbeira, AN et al., Nat. Commun., 2018, Exploring the phenotypic consequences of tissue specific gene expression variation inferred from GWAS summary statistics	29739930	10.1038/s41467-018-03621-1	Cited
Barbeira, AN et al., PLOS Genet., 2019, Integrating predicted transcriptome from multiple tissues improves association detection	30668570	10.1371/journal.pgen.1007889	Cited
Bolger, AM et al., Bioinformatics, 2014, Trimmomatic: A flexible trimmer for Illumina sequence data	24695404	10.1093/bioinformatics/btu170	Cited
Browning, BL et al., Am. J. Hum. Genet., 2018, A one-penny imputed genome from next-generation reference panels	30100085	10.1016/j.ajhg.2018.07.015	Cited
Bulik-Sullivan, BK et al., Nat. Genet., 2015, LD Score regression distinguishes confounding from polygenicity in genome-wide association studies	25642630	10.1038/ng.3211	Cited
Chen, L et al., J. Biol. Chem., 2005, Functionally diverse complement of large conductance calcium- and voltage-activated potassium channel (BK) alpha-subunits generated from a single site of splicing	16081418	10.1074/jbc.M505383200	Cited
Choi, SW et al., Gigascience, 2019, PRSice-2: Polygenic Risk Score software for biobank-scale data	31307061	10.1093/gigascience/giz082	Cited
Choi, SW et al., Nat. Protoc., 2020, Tutorial: A guide to performing polygenic risk score analyses	32709988	10.1038/s41596-020-0353-1	Cited
Dobin, A et al., Bioinformatics, 2013, STAR: Ultrafast universal RNA-seq aligner	23104886	10.1093/bioinformatics/bts635	Cited
Donadoni, M et al., Cell Death Dis., 2019, Alcohol exposure alters pre-mRNA splicing of antiapoptotic Mcl-1L isoform and induces apoptosis in neural progenitors and immature neurons	31171771	10.1038/s41419-019-1673-3	Cited
Farris, SP et al., Mol. Psychiatry, 2015, Transcriptome organization for chronic alcohol abuse in human brain	25450227	10.1038/mp.2014.159	Cited
Gandal, MJ et al., Science, 2018, Shared molecular neuropathology across major psychiatric disorders parallels polygenic overlap	29439242	10.1126/science.aad6469	Cited
Gandal, MJ et al., Science, 2018, Transcriptome-wide isoform-level dysregulation in ASD, schizophrenia, and bipolar disorder	30545856	10.1126/science.aat8127	Cited
Gilpin, NW et al., Biol. Psychiatry, 2015, The central amygdala as an integrative hub for anxiety and alcohol use disorders	25433901	10.1016/j.biopsych.2014.09.008	Cited
Grant, KA et al., Alcohol. Clin. Exp. Res., 2008, Drinking typography established by scheduled induction predicts chronic heavy drinking in a monkey model of ethanol self-administration	18702645	10.1111/j.1530-0277.2008.00765.x	Cited
Heck, J et al., Neuron, 2019, Transient confinement of Ca(V)2.1 Ca(2+)-channel splice variants shapes synaptic short-term plasticity	31104951	10.1016/j.neuron.2019.04.030	Cited
Huggett, SB et al., J. Neurosci., 2020, Genetic architecture and molecular neuropathology of human cocaine addiction	32457073	10.1523/JNEUROSCI.2879-19.2020	Cited
Iancu, OD et al., Addict. Biol., 2018, On the relationships in rhesus macaques between chronic ethanol consumption and the brain transcriptome	28247455	10.1111/adb.12501	Cited
Janak, PH et al., Nature, 2015, From circuits to behaviour in the amygdala	25592533	10.1038/nature14188	Cited
Johnson, EC et al., Lancet Psychiatry, 2020, A large-scale genome-wide association study meta-analysis of cannabis use disorder	33096046	10.1016/S2215-0366(20)30339-4	Primary
Kapoor, M et al., Transl. Psychiatry, 2019, Analysis of whole genome-transcriptomic organization in brain to identify genes associated with alcoholism	30765688	10.1038/s41398-019-0384-y	Primary
Kranzler, HR et al., Nat. Commun., 2019, Genome-wide association study of alcohol consumption and use disorder in 274,424 individuals from multiple populations	30940813	10.1038/s41467-019-09480-8	Primary
Lee, C et al., Biol. Psychiatry, 2014, Altered gamma-aminobutyric acid type B receptor subunit 1 splicing in alcoholics	24209778	10.1016/j.biopsych.2013.08.028	Cited
Li, YI et al., Nat. Commun., 2019, Prioritizing Parkinson's disease genes using population-scale transcriptomic data	30824768	10.1038/s41467-019-08912-9	Cited
Li, YI et al., Nat. Genet., 2018, Annotation-free quantification of RNA splicing using LeafCutter	29229983	10.1038/s41588-017-0004-9	Cited
Liu, M et al., Nat. Genet., 2019, Association studies of up to 1.2 million individuals yield new insights into the genetic etiology of tobacco and alcohol use	30643251	10.1038/s41588-018-0307-5	Primary
Love, MI et al., Genome Biol., 2014, Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2	25516281	10.1186/s13059-014-0550-8	Cited
Palmer, RH et al., Addiction, 2015, Shared additive genetic influences on DSM-IV criteria for alcohol dependence in subjects of European ancestry	26211938	10.1111/add.13070	Cited
Pan, Q et al., Nat. Genet., 2008, Deep surveying of alternative splicing complexity in the human transcriptome by high-throughput sequencing	18978789	10.1038/ng.259	Cited
Pei, W et al., Biochemistry, 2009, Flip and flop: A molecular determinant for AMPA receptor channel opening	19275243	10.1021/bi8015907	Cited
Penn, AC et al., Neuron, 2012, Activity-mediated AMPA receptor remodeling, driven by alternative splicing in the ligand-binding domain	23141062	10.1016/j.neuron.2012.08.010	Cited
Raj, T et al., Nat. Genet., 2018, Integrative transcriptome analyses of the aging brain implicate altered splicing in Alzheimer's disease susceptibility	30297968	10.1038/s41588-018-0238-1	Cited
Rao, X et al., Mol. Psychiatry, 2021, Allele-specific expression and high-throughput reporter assay reveal functional genetic variants associated with alcohol use disorders	31477794	10.1038/s41380-019-0508-z	Primary
Saad, MH et al., Sci. Rep., 2019, Differentially expressed gene netowrks, biomarkers long noncoding RNAs and shared responses with cocaine identified in the midbrains of human opioid abusers	30733491	10.1038/s41598-018-38209-8	Cited
Schlesinger, F et al., Synapse, 2005, Desensitization and resensitization are independently regulated in human recombinant GluR subunit coassemblies	15635696	10.1002/syn.20110	Cited
Shen, S et al., Proc. Natl. Acad. Sci., 2014, rMATS: Robust and flexible detection of differential alternative splicing from replicate RNA-Seq data	25480548	10.1073/pnas.1419161111	Cited
Signor, S et al., Heredity (Edinb.), 2018, Dynamic changes in gene expression and alternative splicing mediate the response to acute alcohol exposure in Drosophila melanogaster	30143789	10.1038/s41437-018-0136-4	Cited
Smedley, D et al., BMC Genomics, 2009, BioMart–biological queries made easy	19144180	10.1186/1471-2164-10-22	Cited
Takata, A et al., Nat. Commun., 2017, Genome-wide identification of splicing QTLs in the human brain and their enrichment among schizophrenia-associated loci	28240266	10.1038/ncomms14519	Cited
Van Booven, D et al., Transl. Psychiatry, 2021, Alcohol use disorder causes global changes in splicing in the human brain	33414398	10.1038/s41398-020-01163-z	Cited
Walter, N et al., Addict. Biol., 2021, Effect of chronic ethanol consumption in rhesus macaques on the nucleus accumbens core transcriptome	33942443	10.1111/adb.13021	Cited
Wang, K et al., Nucleic Acids Res., 2010, ANNOVAR: Functional annotation of genetic variants from high-throughput sequencing data	20601685	10.1093/nar/gkq603	Cited
Zhang, X et al., Nat. Genet., 2015, Identification of common genetic variants controlling transcript isoform variation in human whole blood	25685889	10.1038/ng.3220	Cited
Zhao, H et al., Bioinformatics, 2014, CrossMap: A versatile tool for coordinate conversion between genome assemblies	24351709	10.1093/bioinformatics/btt730	Cited
Zhou, H et al., JAMA Psychiat., 2020, Association of OPRM1 functional coding variant with opioid use disorder: A genome-wide association study	32492095	10.1001/jamapsychiatry.2020.1206	Cited
Zhou, H et al., Nat. Neurosci., 2020, Genome-wide meta-analysis of problematic alcohol use in 435,563 individuals yields insights into biology and relationships with other traits	32451486	10.1038/s41593-020-0643-5	Primary

In this knowledge base

Title	Year	PMID
RNA alternative splicing impacts the risk for alcohol use disorder.	2023	37217680

External

Title	Authors	Journal	Year	Link
Ethanol induces neuroimmune dysregulation and soluble TREM2 generation in a human iPSC neuron, astrocyte, microglia triculture model.	Boreland AJ et al.	—	2026	→
Alternative splicing in addiction.	Bhatnagar A et al.	—	2025	→
Comparative mRNA profile analysis from NAc of adolescent male mice after binge-like alcohol exposure eliciting deficits in context fear extinction learning.	Lloret Torres ME et al.	—	2025	→
Global Changes in Gene Expression and Splicing in Alcoholic Liver Disease	Blokhin IO et al.	—	2025	—
It is not just about transcription: involvement of brain RNA splicing in substance use disorders.	Carvalho L et al.	—	2024	→
Multi-omics profiling of DNA methylation and gene expression alterations in human cocaine use disorder.	Zillich E et al.	—	2024	→
Population-level exploration of alternative splicing and its unique role in controlling agronomic traits of rice.	Zhang H et al.	—	2024	→
The genetical genomic path to understanding why rats and humans consume too much alcohol.	Tabakoff B et al.	—	2024	→
RNA alternative splicing impacts the risk for alcohol use disorder.	Li R et al.	—	2023	→
Integrating human brain proteomic data with genome-wide association study findings identifies novel brain proteins in substance use traits.	Toikumo S et al.	—	2022	→
Opioid Use Disorder and Alternative mRNA Splicing in Reward Circuitry.	Huggett SB et al.	—	2022	→