Exon resequencing of H3K9 methyltransferase complex genes, EHMT1, EHTM2 and WIZ, in Japanese autism subjects.
BACKGROUND
Histone H3 methylation at lysine 9 (H3K9) is a conserved epigenetic signal, mediating heterochromatin formation by trimethylation, and transcriptional silencing by dimethylation. Defective GLP (Ehmt1) and G9a (Ehmt2) histone lysine methyltransferases, involved in mono and dimethylation of H3K9, confer autistic phenotypes and behavioral abnormalities in animal models. Moreover, EHMT1 loss of function results in Kleefstra syndrome, characterized by severe intellectual disability, developmental delays and psychiatric disorders. We examined the possible role of histone methyltransferases in the etiology of autism spectrum disorders (ASD) and suggest that rare functional variants in these genes that regulate H3K9 methylation may be associated with ASD.


METHODS
Since G9a-GLP-Wiz forms a heteromeric methyltransferase complex, all the protein-coding regions and exon/intron boundaries of EHMT1, EHMT2 and WIZ were sequenced in Japanese ASD subjects. The detected variants were prioritized based on novelty and functionality. The expression levels of these genes were tested in blood cells and postmortem brain samples from ASD and control subjects. Expression of EHMT1 and EHMT2 isoforms were determined by digital PCR.


RESULTS
We identified six nonsynonymous variants: three in EHMT1, two in EHMT2 and one in WIZ. Two variants, the EHMT1 ankyrin repeat domain (Lys968Arg) and EHMT2 SET domain (Thr961Ile) variants were present exclusively in cases, but showed no statistically significant association with ASD. The EHMT2 transcript expression was significantly elevated in the peripheral blood cells of ASD when compared with control samples; but not for EHMT1 and WIZ. Gene expression levels of EHMT1, EHMT2 and WIZ in Brodmann area (BA) 9, BA21, BA40 and the dorsal raphe nucleus (DoRN) regions from postmortem brain samples showed no significant changes between ASD and control subjects. Nor did expression levels of EHMT1 and EHMT2 isoforms in the prefrontal cortex differ significantly between ASD and control groups.


CONCLUSIONS
We identified two novel rare missense variants in the EHMT1 and EHMT2 genes of ASD patients. We surmise that these variants alone may not be sufficient to exert a significant effect on ASD pathogenesis. The elevated expression of EHMT2 in the peripheral blood cells may support the notion of a restrictive chromatin state in ASD, similar to schizophrenia.
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Shabeesh
S
Balan
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Yoshimi
Y
Iwayama
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Motoko
M
Maekawa
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Tomoko
T
Toyota
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Tetsuo
T
Ohnishi
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Manabu
M
Toyoshima
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Chie
C
Shimamoto
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Kayoko
K
Esaki
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan.
Kazuo
K
Yamada
Department of Psychiatry and Neurology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan.
Yasuhide
Y
Iwata
Department of Psychiatry and Neurology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan.
Katsuaki
K
Suzuki
Department of Psychiatry, Division of Clinical Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan.
Masayuki
M
Ide
Graduate School of Information Science, Nagoya University, Nagoya, Aichi, Japan.
Motonori
M
Ota
Faculty of Engineering, Maebashi Institute of Technology, Maebashi, Japan.
Satoshi
S
Fukuchi
Faculty of Sociology, Chukyo University, Chukyo, Aichi, Japan ; Research Center for Child Mental Development, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan.
Masatsugu
M
Tsujii
Department of Psychiatry and Neurology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan ; Research Center for Child Mental Development, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan.
Norio
N
Mori
Cellular Memory Laboratory, RIKEN, Wako, Saitama, Japan ; CREST (Core Research for Evolutionary Science and Technology), Japan Science and Technology Agency, Kawaguchi, Saitama, Japan.
Yoichi
Y
Shinkai
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama, 351-0198 Japan ; CREST (Core Research for Evolutionary Science and Technology), Japan Science and Technology Agency, Kawaguchi, Saitama, Japan.
Takeo
T
Yoshikawa
Autism; G9a; GLP; H3K9; Histone methyltransferase; Rare variant; Wiz