DNA Methylation of HTR2A, DRD2 & RELN Genes Promoters and CpG rich areas in
Schizophrenia.
Hamid
Mostafavi Abdolmaleky, M.D.1,2,6, Sam Thiagalingam, Ph.D.3,
Cassandra L. Smith, Ph.D. 4 , Stephen V.Faraone, Ph.D.2,5,
Kunag-hung Cheng, MS. 3 ,Rahim Shafa M.D.7, Stephen Glatt
Ph.D.1,2 , William S Stone, Ph.D. 1,2, and Ming Tsuang,
M.D.1,2,,5,8,9
1Department
of Psychiatry, Harvard Medical School at Massachusetts Mental Health Center
2Harvard
Institute of Psychiatric Epidemiology and Genetics, Boston, Massachusetts
3Department
of Genetics and Genomics, Boston University School of Medicine, Boston, MA
4Laboratory of
Molecular Biotechnology Research, Center for Advanced Biotechnology, and
Departments of Biomedical Engineering, Biology and Pharmacology, Boston
University, Boston, MA.
5Department of
Psychiatry, Harvard Medical School, Massachusetts General Hospital, Boston, MA
6Department
of Psychiatry, Iran University of Medical Sciences, Tehran, Iran
7Metrowest
CNS Research Center, Tenet Metrowest Medical Center, Natic, MA
8Psychiatry
Service, Brockton-West Roxbury Veterans Affairs Medical Center, Brockton,
Massachusetts
9Department
of Epidemiology, Harvard School of Public Health, Boston, Massachusetts
Abstract
Introduction:
Cytosine methylation can prevent gene expression through the actions of four
known proteins, MeCP2, MBD1, MBD2 and MBD3 which bind to methylated DNA.
Concurrently, histones are deacetylated and the
chromatin structure of the chromosome is modified and transcription is reduced.
Methylation can be affected by dietary level of
methyl-donor components, such as methionine and folic acid. Also, the growth of
granule cells in culture medium under sub-optimal conditions resulted in
apoptosis due to a reduction in DNA methylation.
DNA
methylation increases in response to transient ischemia that endangers CNS
neurons survival, while inhibition of DNA methylation prevents brain damages.
Fetal
hypoxia produces more structural brain abnormalities in schizophrenic patients
and their non schizophrenic siblings, especially with low birth weight. There is
evidence indicating that DNA methylation could be abnormal in the RELN, DRD2 and
HTR2A genes in schizophrenia and mood disorder.
Objectives:
Comparing the pattern and amount of DNA Methylation in the promoters and CpG
rich areas of the HTR2A, DRD2 and RELN genes in post-mortem brain tissues of
schizophrenic patients and normal controls.
Materials and methods:
Five samples for each group were provided by the Harvard Brain Tissue Resources
Center. Several primers were specified and tested for detection of the DRD2,
HTR2A and RELN genes’ promoters and CpG rich areas. The technique of direct
sequencing of sodium bisulfite-treated DNA was applied for mapping of
5-methylated cytosine and the technique of Methylation specific PCR was used to
specify the site specific methylation difference among patients and controls.
Results:
Promoter areas of these genes are sparsely methylated, however CpG rich areas
located in the non-promoter (intronic) areas are heavily methylated in patients
and controls. We found some differences between samples and controls in the
promoter area especially in the reelin gene. However, we were unable to achieve
statistical significance due to the small sample size. We have been provided
with an additional 105 samples from the Stanley Medical Research Institute to
overcome the small sample size problems, and have included bipolar disorder in
the project as well.
Key words:
DNA Methylation, Schizophrenia, Bipolar, HTR2A, DRD2, RELN Gene, Frontal lobe,
Post-mortem.