Protein kinase A determines timing of early differentiation through epigenetic regulation with G9a

Kohei Yamamizu; Mayako Fujihara; Makoto Tachibana; Shiori Katayama; Akiko Takahashi; Eiji Hara; Hiroshi Imai; Yoichi Shinkai; Jun K Yamashita

doi:10.1016/j.stem.2012.02.022

Protein kinase A determines timing of early differentiation through epigenetic regulation with G9a

Cell Stem Cell. 2012 Jun 14;10(6):759-770. doi: 10.1016/j.stem.2012.02.022.

Authors

Kohei Yamamizu¹, Mayako Fujihara², Makoto Tachibana³, Shiori Katayama¹, Akiko Takahashi⁴, Eiji Hara⁴, Hiroshi Imai², Yoichi Shinkai³, Jun K Yamashita⁵

Affiliations

¹ Department of Stem Cell Differentiation, Institute for Frontier Medical Sciences, Kyoto University, Kyoto 606-8507, Japan; Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto 606-8507, Japan.
² Laboratory of Reproductive Biology, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan.
³ Experimental Research Center for Infectious Disease, Institute for Virus Research, Kyoto University, Kyoto 606-8507, Japan.
⁴ Cancer Institute, Japanese Foundation for Cancer Research, Tokyo 135-8550, Japan.
⁵ Department of Stem Cell Differentiation, Institute for Frontier Medical Sciences, Kyoto University, Kyoto 606-8507, Japan; Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto 606-8507, Japan. Electronic address: [email protected].

PMID: 22704517
DOI: 10.1016/j.stem.2012.02.022

Abstract

Timing of cell differentiation is strictly controlled and is crucial for normal development and stem cell differentiation. However, underlying mechanisms regulating differentiation timing are fully unknown. Here, we show a molecular mechanism determining differentiation timing from mouse embryonic stem cells (ESCs). Activation of protein kinase A (PKA) modulates differentiation timing to accelerate the appearance of mesoderm and other germ layer cells, reciprocally correlated with the earlier disappearance of pluripotent markers after ESC differentiation. PKA activation increases protein expression of G9a, an H3K9 methyltransferase, along with earlier H3K9 dimethylation and DNA methylation in Oct3/4 and Nanog gene promoters. Deletion of G9a completely abolishes PKA-elicited acceleration of differentiation and epigenetic modification. Furthermore, G9a knockout mice show prolonged expressions of Oct3/4 and Nanog at embryonic day 7.5 and delayed development. In this study, we demonstrate molecular machinery that regulates timing of multilineage differentiation by linking signaling with epigenetics.

Publication types

Research Support, Non-U.S. Gov't
Retracted Publication

MeSH terms

Animals
Blotting, Western
Cell Differentiation*
Chromatin Immunoprecipitation
Cyclic AMP-Dependent Protein Kinases / genetics
Cyclic AMP-Dependent Protein Kinases / metabolism*
DNA Methylation*
Embryo, Mammalian / cytology*
Embryo, Mammalian / metabolism
Embryonic Stem Cells / cytology*
Embryonic Stem Cells / metabolism
Epigenesis, Genetic*
Female
Gene Expression Regulation, Developmental
Histone-Lysine N-Methyltransferase / physiology*
Histones / metabolism
Homeodomain Proteins / genetics
Homeodomain Proteins / metabolism
In Situ Hybridization
Mice
Mice, Knockout
Nanog Homeobox Protein
Octamer Transcription Factor-3 / genetics
Octamer Transcription Factor-3 / metabolism
RNA, Messenger / genetics
Real-Time Polymerase Chain Reaction
Regulatory Sequences, Nucleic Acid / genetics
Reverse Transcriptase Polymerase Chain Reaction
Time Factors

Substances

Histones
Homeodomain Proteins
Nanog Homeobox Protein
Nanog protein, mouse
Octamer Transcription Factor-3
Pou5f1 protein, mouse
RNA, Messenger
G9a protein, mouse
Histone-Lysine N-Methyltransferase
Cyclic AMP-Dependent Protein Kinases