Neuronal nAchR genes in smoking behavior and addiction Open Access

Wiste, Anna Katrina (2009)

Permanent URL: https://etd.library.emory.edu/concern/etds/df65v800k?locale=en
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Abstract

The nicotinic acetylcholine receptor subunit gene cluster on chromosome 15
containing CHRNA5, CHRNA3 and CHRNB4 has been highlighted in several
association studies of genetic influences on smoking and smoking-related diseases. We
reported genome-wide significant association of the synonymous SNP, rs1051730, with
smoking quantity, and also demonstrated that the T allele of this variant carried an
increased risk of nicotine dependence, lung cancer and peripheral arterial disease.
Given the high rate of smoking among individuals with other substance
dependence disorders, we sought to determine whether this variant also increased risk for
dependence on other substances. Subjects met DSM criteria for dependence for at least
one substance of abuse, including alcohol, amphetamine, cannabis, opiates, and
sedatives/anxiolytics. Smoking quantity information was available from questionnaire
data for a subset of subjects. Among alcohol dependent subjects the variant had the same
effect on smoking quantity as observed in the general population.
To characterize the region, the role of variants within the region in regulation
of gene expression was explored. Blood and adipose tissue from a large sample of
subjects was used to test for association of the variant to expression of CHRNA5. Several
correlated variants within the region show significant association to expression of
CHRNA5.
The association with smoking and smoking-related disease reported in our
study and others has been observed in genome-wide association studies. In most cases,
association signals detected in genome-wide association studies are expected to be
indirect, occurring through linkage disequilibrium with a functional variant. To identify
the most likely functional variant responsible for the association signal we have
performed sequencing in the gene cluster in 3 samples: nicotine dependence, lung cancer,
and low quantity smokers. Among common variants in the cluster no variants were
identified with stronger association than rs1051730. A functional SNP in CHRNA5,
rs16969968, is in perfect LD with rs1051730 in Iceland, and it is the only common
functional SNP in the cluster. Based upon these factors, rs16969968 is currently the
variant most likely to be responsible for the association signals observed in this cluster to
smoking behavior and smoking-related diseases.

Table of Contents


Table of Contents
General Introduction…………………………………………………………………1
Chapter 1: Introduction……………..……………………………………………....2

A. Genome-wide Association Studies and Their Impact on Genetics of
Complex Disease......................................................................................................2
B. Definition of Phenotypes.....................................................................................5
C. Genetic Basis of Smoking Behavior and Nicotine Dependence........................12
D. Nicotinic acetylcholine receptors - structure and role in nicotine
Dependence.............................................................................................................12
E. Identification of Risk Variant in CHRNA5/CHRNA3/CHRNB4 gene
cluster......................................................................................................................14
F. Additional findings on the CHRNA5/CHRNA3/CHRNB4 gene cluster
variant......................................................................................................................17
G. Previous Characterizations of the Region...........................................................18
H. Functional Importance of Genes in the Cluster..................................................19
I. Objectives.............................................................................................................20


Chapter 2: Association of Smoking Quantity with a CHRNA5/CHRNA3/CHRNB4
variant in Substance Dependence, but no Association of Variants within the Cluster
to Substance Dependence itself.

A. Abstract………………………………………………………………………...23

B. Introduction………………………………………………………………….....24

C. Materials and Methods...……………………………………………………….25

D. Results………………………………………………………………………….28

E. Discussion……………………………………………………………………..29

Chapter 3: Characterization of the CHRNA5/CHRNA3/CHRNB4 nicotinic
acetylcholine receptor gene cluster in nicotine dependence and lung cancer

A. Abstract………………………………………………………………………...39
B. Introduction…………………………………………………………………….40
C. Results………………………………………………………………………….42
D. Discussion……………………………………………………………………...48
E. Materials and Methods……………………………………………………...….51

Chapter 4 Discussion………………………………………………………..………..74

A. Summary of Results…………………………………………………………….75
B. Other substances of abuse....................................................................................76
C. Sequencing and Expression Analysis...................................................................77
D. Conclusions……………………………………………………………………..80

Chapter 5: References………………………………………………………………..83




Figures and Tables

Figure 1.1 - DSM-IV Criteria for Substance Dependence.......................................10
Figure 1.2 - Fagerström Test of Nicotine Dependence (FTND)…………………...11
Table 2.1 - Demographic Description………………………………………………32
Table 2.2 - Association of Variants in the LD block with Alcohol Dependence....33
Table 2.3 - Association of Variants Tested to Other Substance Dependence

Diagnoses………………………………………………………………..34
Table 2.4 - Regression Analysis of rs1051730 and Smoking Quantity…………...35
Table 2.5.A - Alcohol and Amphetamine Dependence……………………………36
Table 2.5.B - Cannabis, Opioid and Sedative, Anxiolytic and Hypnotic (SAH)

Dependence……………………………………………………………37
Figure 3.1 - Illustration of the risk for nicotine dependence observed for
rs1051730 (1) and rs578776 (2) based on the comparison of 2161 nicotine
dependent individuals and 865 low quantity smokers..............................................57
Figure 3.2 - The short allele of rs3841324 is associated with increased
expression of CHRNA5 RNA in blood and subcutaneous adipose tissue...............58
Table 3.1 - Comparison of frequency of markers with minor allele frequency

> 1% in Lung Cancer (LC), Nicotine Dependence (ND) and Low

Quantity Smokers (LQS)………….........……………………………….59
Table 3.2 - Equivalence classes for SNPs with minor allele frequency greater

than 5%.......................................................................................................61
Table 3.3 - Association Results for Equivalences Classes in Larger Samples…....62
Table 3.4 - Association Results for Insertion/Deletions and Microsatellites….…..63
Table 3.5 - Association analysis of CHRNA5 expression with variants in the

LD block………………………………………………………………….64
Table 3.6 - Demographics: Sequencing Cohort…………………………………….65
Table 3.7 - Descriptive information on all variants from sequencing…………….66
Table 3.8 - All variants with frequency less than 1%...............................................69
Table 3.9 - Demographics for cohorts used in analyses including additional

genotyping……………………………………………………………….70
Table 3.10 - Full list of sequencing primers used………………………………….71
Table 3.11 - Build 36 positions for regions sequenced…………………………….74








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