AIMS

Specific Aim 1: Longitudinal methylomic analyses in each trajectory group

o Perform a hypothesis-free genome-wide screen of DNA methylation in the 200 individuals using the Illumina Infinium 27K Methylation array on bisulfite-treated DNA from samples obtained at birth, age 7 and age 15

o Perform extended fine-mapping verification analyses across specific genomic regions nominated from the microarray screen using Pyrosequening and/or the Sequenom EpiTYPER system in larger numbers of cases and controls.

o Assess the extent to which dynamic patterns of methylation vary across the trajectories

o Is there variation in the epigenetic-regulation of genes in the trajectories relative to controls?

? We would estimate "methylation trajectories" conditional on the conduct trajectories in an attempt to sort "noise" from the potentially informative changes in gene regulation

Specific Aim 2: Examine candidate-gene DNA methylation patterns

o Focus on HPA axis regulatory genes known to functionally inter-relate with serotonin

o e.g., corticotropin-releasing hormone (CRH), CRH receptors, CRH binding proteins, pro-opiomelanocortin (POMC), steroid hormone biosynthetic enzymes, etc.).

o Focus on testosterone (androgen) receptors, androgen biosynthesis pathways, monoamine/catecholamine-related genes (e.g., monoamine oxidase, transporters)

o Quantitative analysis of DNA methylation across gene promoter regions using using Pyrosequening and/or the Sequenom EpiTYPER system .

o Perform extended fine-mapping verification analyses across specific genomic regions nominated from the microarray screen using Pyrosequening and/or the Sequenom EpiTYPER system in cases and controls.

o Assess the extent to which dynamic patterns of methylation vary across the trajectories

o Is there variation in the epigenetic regulation of genes in the trajectories relative to controls?

We would estimate "methylation trajectories" conditional on the conduct trajectories in an attempt to sort "noise" from the potentially informative changes in regulation

Specific Aim 3: examine associations between DNA methylation trajectories and environmental

risk exposures from gestation though to adolescence

3.1 Data collection.

DNA from samples obtained at birth, age 7 and age 15

3.2 Existing Data Required

Environmental risk data are listed in the previous section. The genes we propose to genotype for the candidate methylation scans are listed in the next section.

3.4 Data Analysis.

See specific aims (above). Epigentic analysis will take place at the The Institute of Psychiatry Psychiatric Epigenetics Group, King's College London.