PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-7 (7)
 

Clipboard (0)
None
Journals
Year of Publication
Document Types
1.  Multiple regulatory variants modulate expression of 5-hydroxytryptamine 2A receptors in human cortex 
Biological psychiatry  2012;73(6):546-554.
Background
The 5-hydroxytryptamine 2A receptor, encoded by HTR2A, is a major post-synaptic target for serotonin in the human brain and a therapeutic drug target. Despite hundreds of genetic associations investigating HTR2A polymorphisms in neuropsychiatric disorders and therapies, the role of genetic HTR2A variability in health and disease remains uncertain.
Methods
To discover and characterize regulatory HTR2A variants, we sequenced whole transcriptomes from ten human brain regions with massively-parallel RNA sequencing and measured allelic expression of multiple HTR2A mRNA transcript variants. Following discovery of functional variants, we further characterized their impact on genetic expression in vitro.
Results
Three polymorphisms modulate the use of novel alternative exons and untranslated regions (UTRs), changing expression of RNA and protein. The frequent promoter variant rs6311, widely implicated in human neuropsychiatric disorders, decreases usage of an upstream transcription start site encoding a longer 5′UTR with greater translation efficiency. rs76665058, located in an extended 3′UTR and unique to individuals of African descent, modulates allelic HTR2A mRNA expression. The third SNP, unannotated and present in only a single subject, directs alternative splicing of exon 2. Targeted analysis of HTR2A in the Sequenced Treatment Alternatives to Relieve Depression (STAR*D) study reveals associations between functional variants and depression severity or citalopram response.
Conclusions
Regulatory polymorphisms modulate HTR2A mRNA expression in an isoform-specific manner, directing the usage of novel untranslated regions and alternative exons. These results provide a foundation for delineating the role of HTR2A and serotonin signaling in CNS disorders.
doi:10.1016/j.biopsych.2012.09.028
PMCID: PMC3582836  PMID: 23158458
serotonin; 5-HT2A; HTR2A; schizophrenia; depression; mRNA expression
2.  Profiling Solute Carrier Transporters in the Human Blood-Brain Barrier 
doi:10.1038/clpt.2013.175
PMCID: PMC3906042  PMID: 24013810
Blood-brain barrier; solute carrier transporter; drug transporter; OCT3; organic cation transporter; MATE1; multidrug and toxin extrusion protein; expression profiling
3.  Cholesteryl Ester Transfer Protein (CETP) Polymorphisms Affect mRNA Splicing, HDL Levels, and Sex-Dependent Cardiovascular Risk 
PLoS ONE  2012;7(3):e31930.
Polymorphisms in and around the Cholesteryl Ester Transfer Protein (CETP) gene have been associated with HDL levels, risk for coronary artery disease (CAD), and response to therapy. The mechanism of action of these polymorphisms has yet to be defined. We used mRNA allelic expression and splice isoform measurements in human liver tissues to identify the genetic variants affecting CETP levels. Allelic CETP mRNA expression ratios in 56 human livers were strongly associated with several variants 2.5–7 kb upstream of the transcription start site (e.g., rs247616 p = 6.4×10−5, allele frequency 33%). In addition, a common alternatively spliced CETP isoform lacking exon 9 (Δ9), has been shown to prevent CETP secretion in a dominant-negative manner. The Δ 9 expression ranged from 10 to 48% of total CETP mRNA in 94 livers. Increased formation of this isoform was exclusively associated with an exon 9 polymorphism rs5883-C>T (p = 6.8×10−10) and intron 8 polymorphism rs9930761-T>C (5.6×10−8) (in high linkage disequilibrium with allele frequencies 6–7%). rs9930761 changes a key splicing branch point nucleotide in intron 8, while rs5883 alters an exonic splicing enhancer sequence in exon 9.
The effect of these polymorphisms was evaluated in two clinical studies. In the Whitehall II study of 4745 subjects, both rs247616 and rs5883T/rs9930761C were independently associated with increased HDL-C levels in males with similar effect size (rs247616 p = 9.6×10−28 and rs5883 p = 8.6×10−10, adjusted for rs247616). In an independent multiethnic US cohort of hypertensive subjects with CAD (INVEST-GENE), rs5883T/rs9930761C alone were significantly associated with increased incidence of MI, stroke, and all-cause mortality in males (rs5883: OR 2.36 (CI 1.29–4.30), p = 0.005, n = 866). These variants did not reach significance in females in either study. Similar to earlier results linking low CETP activity with poor outcomes in males, our results suggest genetic, sex-dependent CETP splicing effects on cardiovascular risk by a mechanism independent of circulating HDL-C levels.
doi:10.1371/journal.pone.0031930
PMCID: PMC3293889  PMID: 22403620
4.  Nicotinic α5 receptor subunit mRNA expression is associated with distant 5′ upstream polymorphisms 
CHRNA5, encoding the nicotinic α5 subunit, is implicated in multiple disorders, including nicotine addiction and lung cancer. Previous studies demonstrate significant associations between promoter polymorphisms and CHRNA5 mRNA expression, but the responsible sequence variants remain uncertain. To search for cis-regulatory variants, we measured allele-specific mRNA expression of CHRNA5 in human prefrontal cortex autopsy tissues and scanned the CHRNA5 locus for regulatory variants. A cluster of six frequent single nucleotide polymorphisms (rs1979905, rs1979906, rs1979907, rs880395, rs905740, and rs7164030), in complete linkage disequilibrium, fully account for a >2.5-fold allelic expression difference and a fourfold increase in overall CHRNA5 mRNA expression. This proposed enhancer region resides more than 13 kilobases upstream of the CHRNA5 transcription start site. The same upstream variants failed to affect CHRNA5 mRNA expression in peripheral blood lymphocytes, indicating tissue-specific gene regulation. Other promoter polymorphisms were also correlated with overall CHRNA5 mRNA expression in the brain, but were inconsistent with allelic mRNA expression ratios, a robust and proximate measure of cis-regulatory variants. The enhancer region and the nonsynonymous polymorphism rs16969968 generate three main haplotypes that alter the risk of developing nicotine dependence. Ethnic differences in linkage disequilibrium across the CHRNA5 locus require consideration of the upstream enhancer variants when testing clinical associations.
doi:10.1038/ejhg.2010.120
PMCID: PMC2995013  PMID: 20700147
Nicotinic receptor; alpha5 subunit; gene expression; nicotine dependence; lung cancer; enhancer
5.  Polymorphisms affecting gene transcription and mRNA processing in pharmacogenetic candidate genes: detection through allelic expression imbalance in human target tissues 
Pharmacogenetics and genomics  2008;18(9):781-791.
Genetic variation in mRNA expression plays a critical role in human phenotypic diversity, but it has proven difficult to detect regulatory polymorphisms - mostly single nucleotide polymorphisms (rSNPs). Additionally, variants in the transcribed region, termed here ‘structural RNA SNPs’ (srSNPs), can affect mRNA processing and turnover. Both rSNPs and srSNPs cause allelic mRNA expression imbalance (AEI) in heterozygous individuals. We have applied a rapid and accurate AEI methodology for testing 42 genes implicated in human diseases and drug response, specifically cardiovascular and CNS diseases, and affecting drug metabolism and transport. Each gene was analyzed in physiologically relevant human autopsy tissues, including brain, heart, liver, intestines, and lymphocytes. Substantial AEI was observed in ∼55% of the surveyed genes. Focusing on cardiovascular candidate genes in human hearts, AEI analysis revealed frequent cis-acting regulatory factors in SOD2 and ACE mRNA expression, having potential clinical significance. SNP scanning to locate regulatory polymorphisms in a number of genes failed to support several previously proposed promoter SNPs discovered with use of reporter gene assays in heterologous tissues, while srSNPs appear more frequent than expected. Computational analysis of mRNA folding indicates that ∼90% of srSNPs affects mRNA folding, and hence potentially function. Our results indicate that both rSNPs and srSNPs represent a still largely untapped reservoir of variants that contribute to human phenotypic diversity.
doi:10.1097/FPC.0b013e3283050107
PMCID: PMC2779843  PMID: 18698231
6.  Financial and Psychological Risk Attitudes Associated with Two Single Nucleotide Polymorphisms in the Nicotine Receptor (CHRNA4) Gene 
PLoS ONE  2009;4(8):e6704.
With recent advances in understanding of the neuroscience of risk taking, attention is now turning to genetic factors that may contribute to individual heterogeneity in risk attitudes. In this paper we test for genetic associations with risk attitude measures derived from both the psychology and economics literature. To develop a long-term prospective study, we first evaluate both types of risk attitudes and find that the economic and psychological measures are poorly correlated, suggesting that different genetic factors may underlie human response to risk faced in different behavioral domains. We then examine polymorphisms in a spectrum of candidate genes that affect neurotransmitter systems influencing dopamine regulation or are thought to be associated with risk attitudes or impulsive disorders. Analysis of the genotyping data identified two single nucleotide polymorphisms (SNPs) in the gene encoding the alpha 4 nicotine receptor (CHRNA4, rs4603829 and rs4522666) that are significantly associated with harm avoidance, a risk attitude measurement drawn from the psychology literature. Novelty seeking, another risk attitude measure from the psychology literature, is associated with several COMT (catechol-O-methyl transferase) SNPs while economic risk attitude measures are associated with several VMAT2 (vesicular monoamine transporter) SNPs, but the significance of these associations did not withstand statistical adjustment for multiple testing and requires larger cohorts. These exploratory results provide a starting point for understanding the genetic basis of risk attitudes by considering the range of methods available for measuring risk attitudes and by searching beyond the traditional direct focus on dopamine and serotonin receptor and transporter genes.
doi:10.1371/journal.pone.0006704
PMCID: PMC2724734  PMID: 19693267
7.  Highly variable mRNA expression and splicing of L-type voltage-dependent calcium channel alpha subunit 1C in human heart tissues 
Pharmacogenetics and genomics  2006;16(10):735-745.
Objectives
The voltage-dependent L-type calcium channel α-subunit 1c (Cav1.2, CACNA1C) undergoes extensive mRNA splicing, leading to numerous isoforms with different functions. L-type calcium channel blockers are used in the treatment of hypertension and arrhythmias, but response varies between individuals. We have studied the interindividual variability in mRNA expression and splicing of CACNA1C, in 65 heart tissue samples, taken from heart transplant recipients.
Methods
Splice variants were measured quantitatively by polymerase chain reaction in 12 splicing loci of CACNA1C mRNA. To search for functional cis-acting polymorphisms, we determined allelic expression ratios for total CACNA1C mRNA and several splice variants using marker single nucleotide polymorphisms in exon 4 and exon 30.
Results
Total CACNA1C mRNA levels varied ∼50-fold. Substantial splicing occurred in six loci generating two or more splice variants, some with known functional differences. Splice patterns varied broadly between individuals. Two heart tissues expressed predominantly the dihydropyridine-sensitive smooth muscle isoform of CACNA1C (containing exon 8), rather than the cardiac isoform (containing exon 8a). Lack of significant allelic expression imbalance, observed with total mRNA and several splice variants, argued against CACNA1C polymorphisms as a cause of variability. Taken together, highly variable splicing can cause profound phenotypic variations of CACNA1C function, potentially associated with disease susceptibility and response to L-type calcium channel blockers.
doi:10.1097/01.fpc.0000230119.34205.8a
PMCID: PMC2688811  PMID: 17001293
cis-acting polymorphism; L-type calcium channel α-subunit 1c; mRNA splicing

Results 1-7 (7)