Promoter-wide hypermethylation of the ribosomal RNA gene promoter in the suicide brain - PubMed (original) (raw)
Promoter-wide hypermethylation of the ribosomal RNA gene promoter in the suicide brain
Patrick O McGowan et al. PLoS One. 2008.
Abstract
Background: Alterations in gene expression in the suicide brain have been reported and for several genes DNA methylation as an epigenetic regulator is thought to play a role. rRNA genes, that encode ribosomal RNA, are the backbone of the protein synthesis machinery and levels of rRNA gene promoter methylation determine rRNA transcription.
Methodology/principal findings: We test here by sodium bisulfite mapping of the rRNA promoter and quantitative real-time PCR of rRNA expression the hypothesis that epigenetic differences in critical loci in the brain are involved in the pathophysiology of suicide. Suicide subjects in this study were selected for a history of early childhood neglect/abuse, which is associated with decreased hippocampal volume and cognitive impairments. rRNA was significantly hypermethylated throughout the promoter and 5' regulatory region in the brain of suicide subjects, consistent with reduced rRNA expression in the hippocampus. This difference in rRNA methylation was not evident in the cerebellum and occurred in the absence of genome-wide changes in methylation, as assessed by nearest neighbor.
Conclusions/significance: This is the first study to show aberrant regulation of the protein synthesis machinery in the suicide brain. The data implicate the epigenetic modulation of rRNA in the pathophysiology of suicide.
Conflict of interest statement
Competing Interests: The authors have declared that no competing interests exist.
Figures
Figure 1. Genotyping of the rRNA promoter.
The rRNA promoter sequence was identical for all suicide subjects and controls. The sequence derived from genotyping is shown above the published rRNA sequence, indicating consensus sequences for primers used for sodium bisulfite mapping (underline) and CpG dinucleotides (bold font), with locations marked relative to the transcription start site (arrow). Differences with the published rRNA sequence, U13369, are highlighted in grey, and the base pair length of each sequence is listed on the right side.
Figure 2. Sodium bisulfite mapping of the rRNA promoter in suicide subjects and controls.
Twenty clones were sequenced for each suicide subject (left side) and control (right side), from 2 to 3 independent PCR reactions. Each line represents one clone. Circles representing CpG dinucleotides follow the 5′ to 3′ order of the rRNA promoter sequence for methylated CpG dinucleotides (filled circles), and unmethylated CpG dinucleotides (open circles).
Figure 3. Hypermethylation of the rRNA promoter in suicide subjects relative to controls.
(A) (above) Vertical lines indicate locations of CpG dinucleotides on the rRNA promoter relative to the transcription start site, indicated with the solid arrow, with primer pairs used for bisulfite mapping marked by dashed arrows. (below) Average percentage of methylation for each CpG site, for suicide subjects (N = 13; black bars) and controls (N = 11; white bars). Data are expressed as mean ± S.E.M. *, P<0.05; **, P<0.01; ***, P<0.001, measured by ANOVA. (B) Multiple regression analysis of the number of methylated CpGs per clone and the number of clones shows a significant interaction between suicide subjects (20 clones × 13 subjects, N = 260 total clones; filled circles) and controls (20 clones × 11 subjects, N = 220 total clones; open circles). There are 26 circles per group, as clones are grouped according to methylation status.
Figure 4. Site-independent hypermethylation of the rRNA promoter in suicide subjects.
Positive correlation between suicide and control rRNA promoter methylation percentage across CpG sites (N = 26), showing a conserved hypermethylation in suicide subjects throughout the promoter region. Each data point is labeled with the position of each CpG dinucleotide in the rRNA promoter, relative to the transcription start site.
Figure 5. Anatomical and Genomic specificity of rRNA hypermethylation.
Average percentage of rRNA promoter methylation for selected subjects with large methylation differences in the hippocampus (A) and in the cerebellum (B) of suicide subjects (N = 4, black bars) and controls (N = 4, white bars) for the same subjects. Data are expressed as mean ± S.E.M. **, P<0.01, measured by unpaired t-test. (C) Multiple regression analysis shows a similar negative relationship between the number of methylated CpGs per clone and the number of clones in cerebellum samples from suicide subjects (20 clones × 4 subjects, N = 80 total clones; filled circles) and controls (20 clones × 4 subjects, N = 80 total clones; open circles). There are 26 circles per group, as clones are grouped according to methylation status. (D) (above) Representative images of genome-wide methylation in the hippocampus for a suicide subject and a control, showing cytosine (C) and 5-methylcytosine (5 mC) content used for nearest neighbor analysis. (below) Quantification of the percentage of methylcytosine, following the formula: [(5-methylcytosine) × 100]/(5-methylcytosine + cytosine), shows no difference between suicide subjects (N = 13, black bar) and controls (N = 11, white bar) in genome-wide levels of methylation (P>0.05), measured by unpaired t-test.
Figure 6. rRNA expression is downregulated in suicide.
(A) Suicide subjects (N = 16, black bar) showed significantly less rRNA expression than controls (N = 9, white bar); Data expressed as mean ± S.E.M. *, P<0.05, measured by unpaired t-test. (B) Among subjects where both methylation and expression of rRNA were analyzed, the correlation between rRNA promoter methylation percentage and expression showed a trend for an inverse relationship between methylation percentage and expression of rRNA in suicide subjects (N = 11, filled circles) and controls (N = 8, open circles).
References
- Turecki G. Suicidal behavior: is there a genetic predisposition? Bipolar Disord. 2001;3:335–349. - PubMed
- Mann JJ. A current perspective of suicide and attempted suicide. Ann Intern Med. 2002;136:302–311. - PubMed
- Roy A, Segal NL, Sarchapone M. Attempted suicide among living co-twins of twin suicide victims. Am J Psychiatry. 1995;152:1075–1076. - PubMed
- Roy A, Segal NL, Centerwall BS, Robinette CD. Suicide in twins. Arch Gen Psychiatry. 1991;48:29–32. - PubMed
- Seguin M, Lesage A, Turecki G, Bouchard M, Chawky N, et al. Life trajectories and burden of adversity: mapping the developmental profiles of suicide mortality. Psychol Med. 2007;37:1575–1583. - PubMed
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Other Literature Sources
Medical