Developmental epigenetic modification regulates stochastic expression of clustered protocadherin genes, generating single neuron diversity.
In the brain, enormous numbers of neurons have functional individuality and distinct circuit specificities. Clustered Protocadherins (Pcdhs), diversified cell-surface proteins, are stochastically expressed by alternative promoter choice and affect dendritic arborization in individual neurons. Here we found that the Pcdh promoters are differentially methylated by the de novo DNA methyltransferase Dnmt3b during early embryogenesis. To determine this methylation's role in neurons, we produced chimeric mice from Dnmt3b-deficient induced pluripotent stem cells (iPSCs). Single-cell expression analysis revealed that individual Dnmt3b-deficient Purkinje cells expressed increased numbers of Pcdh isoforms; in vivo, they exhibited abnormal dendritic arborization. These results indicate that DNA methylation by Dnmt3b at early embryonic stages regulates the probability of expression for the stochastically expressed Pcdh isoforms. They also suggest a mechanism for a rare human recessive disease, the ICF (Immunodeficiency, Centromere instability, and Facial anomalies) syndrome, which is caused by Dnmt3b mutations.
KOKORO-Biology Group, Laboratories for Integrated Biology, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Shunsuke
S
Toyoda
KOKORO-Biology Group, Laboratories for Integrated Biology, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Masahumi
M
Kawaguchi
Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan; Japan Science Technology Agency, ERATO, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
Toshihiro
T
Kobayashi
Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Division of Developmental Neurophysiology, National Institute for Physiological Sciences, National Institutes of Natural Sciences, Okazaki, Aichi 444-8787, Japan.
Etsuko
E
Tarusawa
KOKORO-Biology Group, Laboratories for Integrated Biology, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Tomoko
T
Toyama
Laboratory for Mammalian Epigenetic Studies, RIKEN Center for Developmental Biology, Minatojima-minamimachi 2-2-3, Chuo-ku, Kobe, Hyogo 650-0047, Japan.
Masaki
M
Okano
Laboratory for Mammalian Epigenetic Studies, RIKEN Center for Developmental Biology, Minatojima-minamimachi 2-2-3, Chuo-ku, Kobe, Hyogo 650-0047, Japan.
Masaaki
M
Oda
Division of Stem Cell Therapy, Center for Stem Cell Biology and Regenerative Medicine, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan; Japan Science Technology Agency, ERATO, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
Hiromitsu
H
Nakauchi
Division of Developmental Neurophysiology, National Institute for Physiological Sciences, National Institutes of Natural Sciences, Okazaki, Aichi 444-8787, Japan; Department of Physiological Sciences, The Graduate University for Advanced Studies, Okazaki, Aichi 444-8585, Japan.
Yumiko
Y
Yoshimura
Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Section of Mammalian Transgenesis, Center for Genetic Analysis of Behavior, National Institute for Physiological Sciences, Okazaki, Aichi 444-8585, Japan.
Makoto
M
Sanbo
Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Department of Physiological Sciences, The Graduate University for Advanced Studies, Okazaki, Aichi 444-8585, Japan; Section of Mammalian Transgenesis, Center for Genetic Analysis of Behavior, National Institute for Physiological Sciences, Okazaki, Aichi 444-8585, Japan.
Masumi
M
Hirabayashi
KOKORO-Biology Group, Laboratories for Integrated Biology, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Teruyoshi
T
Hirayama
KOKORO-Biology Group, Laboratories for Integrated Biology, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Takahiro
T
Hirabayashi
KOKORO-Biology Group, Laboratories for Integrated Biology, Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan; Japan Science and Technology Agency-Core Research for Evolutional Science and Technology, CREST, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan. Electronic address: yagi@fbs.osaka-u.ac.jp.
Takeshi
T
Yagi
D000367:Age Factors; D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000166:cytology / Q000196:embryology / Q000254:growth & development / Q000378:metabolism; D015820:Cadherins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000172:deficiency / Q000235:genetics; D019175:DNA Methylation; D004622:Embryo, Mammalian; D044127:Epigenesis, Genetic / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental / Q000235:genetics / Q000502:physiology*; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D005810:Multigene Family / Q000235:genetics*; D009474:Neurons / Q000502:physiology*; D039904:Pluripotent Stem Cells / Q000502:physiology; D011401:Promoter Regions, Genetic / Q000502:physiology*; D013269:Stochastic Processes; D000097584:DNA Methyltransferase 3B
D015820:Cadherins; D004248:DNA (Cytosine-5-)-Methyltransferases; D000097584:DNA Methyltransferase 3B