The serotonin reuptake transporter (5HTT) is thought to be the principal regulator of serotonergic activity and epigenetic effects at this locus are thought to be important moderators of vulnerability to neuropsychiatric illness. In attempt to understand the basis of this regulation, several gene polymorphisms that affect 5HTT mRNA levels have been described. But to date, no clear mechanism linking these polymorphisms to vulnerability to epigenetic effects have been described. In this communication, we describe a CpG island in the 5 0 region of the 5HTT gene that contains an alternative exon 1 and possible promoter for 5HTT. We then confirm the existence of this transcript and ascertain the methylation status of this CpG island in 49 lymphoblast cell lines and analyze the relationship between methylation and 5HTT mRNA levels. We demonstrate that methylation at this CpG island is associated with decreased levels of 5HTT mRNA, but that this effect is evident only when 5HTTLPR genotype is taken into account. We suggest that these findings have significant implications for the understanding of the role of this locus in behavioral illness. INTRODUCTIONThe serotonin transporter (5HTT) located at 17q11 is thought to be a major regulator of serotonergic neurotransmission and thereby vulnerability to behavioral illness. In efforts to determine whether genetic variation at 5HTT contributes variation in serotonin reuptake, a large number of studies have been conducted. To a large extent, these studies have failed to demonstrate a significant role for coding variants in regulating serotonin re-uptake. In contrast, several studies have demonstrated that non-coding variants have a significant role in regulating serotonin transport by altering 5HTT mRNA levels. The best characterized of these non-coding variants, the 5HTTLPR polymorphism, has effects on 5HTT mRNA levels in lymphoblasts Bradley et al., 2005], in platelet serotonin reuptake [Greenberg et al., 1999;Nobile et al., 1999] and platelet serotonin content [Hanna et al., 1998]. Other non-coding variants including the recently reported Long G polymorphism [Hu et al., 2005] and an intron 2 polymorphism denoted as the STin2 variable number tandem repeat (VNTR) [Hranilovic et al., 2004] also may have effects on serotonin mRNA production, although these effects are less well established.Given the importance of this gene in regulating serotonergic neurotransmission, at least several hundred studies have examined the role of the genetic variation in 5HTT, in particular, the 5HTTLPR polymorphism, in mediating vulnerability to neuropsychiatric illness such as depression, schizophrenia, and autism. The results of these studies have been mixed, with many studies inferring that the 5HTTLPR polymorphism may have effect on vulnerability to a broad range of behavioral illnesses [Conroy et al., 2004;Gorwood, 2004;Lotrich and Pollock, 2004;Savitz and Ramesar, 2004]. However, with the exception of alcoholism and schizophrenia [Fan and Sklar, 2005;Lasky-Su et al., 2005], meta-analyses of...

Substance abuse has an enormous impact on economic and quality of life measures throughout the world. In more developed countries, overutilization of the most common forms of substances of abuse, alcohol and tobacco, is addressed primarily through prevention of substance use initiation and secondarily through the treatment of those with substance abuse or dependence. In general, these therapeutic approaches to substance abuse are deemed effective. However, there is a broad consensus that the development of additional tools to aid diagnosis, prioritize treatment selection and monitor treatment response could have substantial impact on the effectiveness of both substance use prevention and treatment. The recent demonstrations by a number of groups that substance use exposure is associated with robust changes in DNA methylation signatures of peripheral blood cells suggests the possibility that methylation assessments of blood or saliva could find broad clinical applications. In this article, we review recent progress in epigenetic approaches to substance use assessment with a particular emphasis on smoking (and alcohol) related applications. In addition, we highlight areas, such as the epigenetics of psychostimulant, opioid and cannabis abuse, which are markedly understudied and could benefit from intensified collaborative efforts to define epigenetic biomarkers of abuse and dependence.

These results suggest that shared genetic susceptibility contributes modestly to MDD and AD comorbidity. Individuals with elevated polygenic risk for MDD may also be at risk for AD.

Better biomarkers to detect smoking are needed given the tremendous public health burden caused by smoking. Current biomarkers to detect smoking have significant limitations, notably a short half-life for detection and lack of sensitivity for light smokers. These limitations may be particularly problematic in populations with less accurate self-reporting. Prior epigenome-wide association studies indicate that methylation status at cg05575921, a CpG residue located in the aryl hydrocarbon receptor repressor (AHRR) gene, may be a robust indicator of smoking status in individuals with as little as half of a pack-year of smoking. In this study, we show that a novel droplet digital PCR assay for measuring methylation at cg05575921 can reliably detect smoking status, as confirmed by serum cotinine, in populations with different demographic characteristics, smoking histories, and rates of false-negative self-report of smoking behavior. Using logistic regression models, we show that obtaining maximum accuracy in predicting smoking status depends on appropriately weighting self-report and cg05575921 methylation according to the characteristics of the sample being tested. Furthermore, models using only cg05575921 methylation to predict smoking perform nearly as well as those also including self-report across populations. In conclusion, cg05575921 has significant potential as a clinical biomarker to detect smoking in populations with varying rates of accuracy in self-report of smoking behavior.

A number of studies have demonstrated that genetic variation at GABRA2 alters vulnerability to alcohol dependence. However, the exact identity of the causal variant(s), the relationship of these variants to other forms of substance use and behavioral illness is uncertain. Therefore, we genotyped 516 subjects from the Iowa Adoption Studies, a large longitudinal case and control adoption study of substance use, at 39 single nucleotide polymorphisms encompassing the GABRA2 locus and analyzed them with respect to their lifetime history of three common forms of substance use (alcohol (AD), nicotine (ND) and cannabis dependence (CD)), major depressive disorder (MDD) and antisocial personality disorder (ASPD) and relevant exposure variables. Using regression analysis, we found substantial evidence that both GABRA2 genotype and haplotype are significantly related to vulnerability to AD, ND and CD with the strongest relationships noted with respect to ND. Consistent with prior studies suggesting exposure is an important step in the development of substance use, we found the inclusion of substance exposure data in to our analytic models markedly increased the strength of the genetic associations of GABRA2 haplotype with substance use. Finally, we report that the genetic effects were markedly more pronounced in females than in males. We conclude that genetic variation at or near the GABRA2 locus significantly effects vulnerability not only to AD, but to other forms of substance use including ND and CD, and that the effects may be sex dependent.