Association and cis-mQTL analysis of variants in CHRNA3-A5, CHRNA7, CHRNB2, and CHRNB4 in relation to nicotine dependence in a Chinese Han population

Abstract

Nicotine dependence (ND) is a worldwide health problem. Numerous genetic studies have demonstrated a significant association of variants in nicotinic acetylcholine receptors (nAChRs) with smoking behaviors. However, most of these studies enrolled only subjects of European or African ancestry. In addition, although an increasing body of evidence implies a causal connection of single-nucleotide polymorphisms (SNPs) and epigenetic regulation of gene expression, few studies of this issue have been reported. In this study, we performed both association and interaction analysis for 67 SNPs in CHRNA3-A5, CHRNA7, CHRNB2, and CHRNB4 with ND in a Chinese Han population (N = 5055). We further analyzed cis-mQTL for the three most significant SNPs and 5580 potential methylation loci within these target gene regions. Our results indicated that the SNPs rs1948 and rs7178270 in CHRNB4 and rs3743075 in CHRNA3 were significantly associated with the Fagerström Test for Nicotine Dependence (FTND) score (p = 6.6 × 10−5; p = 2.0 × 10−4, and p = 7.0 × 10−4, respectively). Haplotype-based association analysis revealed that two major haplotypes, T-G and C-A, formed by rs3743075–rs3743074 in CHRNA3, and other two major haplotypes, A-G-C and G-C-C, formed by rs1948–rs7178270–rs17487223 in CHRNB4, were significantly associated with the FTND score (p ≤ 8.0 × 10−4). Further, we found evidence for the presence of significant interaction among variants within CHRNA3/B4/A5, CHRNA4/B2/A5, and CHRNA7 in affecting ND, with corresponding p values of 5.8 × 10−6, 8.0 × 10−5, and 0.012, respectively. Finally, we identified two CpG sites (CpG_2975 and CpG_3007) in CHRNA3 that are significantly associated with three cis-mQTL SNPs (rs1948, rs7178270, rs3743075) in the CHRNA5/A3/B4 cluster (p ≤ 1.9 × 10−6), which formed four significant CpG–SNP pairs in our sample. Together, we revealed at least three novel SNPs in CHRNA3 and CHRNB4 to be significantly associated with the FTND score. Further, we showed that these significant variants contribute to ND via two methylated sites, and we demonstrated significant interaction affecting ND among variants in CHRNA5/A3/B4, CHRNA7, and CHRNA4/B2/A5. In sum, these findings provide robust evidence that SNPs in nAChR genes convey a risk of ND in the Chinese Han population.

Association and cis-mQTL analysis of variants in serotonergic genes associated with nicotine dependence in Chinese Han smokers

“…By using a dataset with all CHRNA5 methylation, expression, and risk of SNPs data available, Hancock et al first revealed that several genetic variations underlying CHRNA5 methylation in the human brain contributed to the risk of ND 60 . Recently, we linked SNP, cis -mQTL, and cis -eQTL 45 and revealed two methylated sites associated significantly with three SNPs forming four significant CpG–SNP pairs in a Chinese Han population. In the present study, as shown in Fig.…”

“…The threshold for the genotyping rate of all SNPs was set as ≥95%, and the p value for the HWE was set as no ≥5%. All procedures and conditions used in this study were essentially the same as what we reported previously on the same samples 44 , 45 .…”

“…The DNA methylation dataset used here consisted of samples from 36 smokers and 36 nonsmokers selected from the sample set described above on the basis of the age, sex, and smoking status, which is an ongoing whole-genome bisulfite sequencing project in this laboratory 45 . Briefly, the Illumina HiSeq X Ten platform was applied to conduct DNA methylation sequencing with an average of about 700 million (±75 million; SD) 150-bp paired-end reads per sample.…”

“…We conducted cis -mQTL analysis with a similar procedure, as described before 45 . The region for the cis- mQTL analysis was set within 50 kb upstream and downstream of each SNP of interest 52 , 53 .…”

Genome-Wide Association Study of Smoking Behavior Traits in a Chinese Han Population

“…To replicate the GWAS associations, 1,329 participants including 805 smokers and 524 nonsmokers were recruited from local hospitals in Jincheng and Taiyuan of Shanxi Province in China during 2012-2013 (12). All 1,329 participants were males, aged 19-62 years (Supplementary Table 1).…”