Dnmt1 (DNA methyltansferase 1) is an enzyme that recognizes and methylates hemimethylated DNA during DNA replication to maintain methylation patterns. The N-terminal region of Dnmt1 is known to form an independent domain structure that interacts with various regulatory proteins and DNA. In the present study, we investigated protein kinases in the mouse brain that could bind and phosphorylate the N-terminal regulatory domain of Dnmt1. A protein fraction containing protein kinase activity for phosphorylation of Dnmt1(1–290) was prepared using Dnmt1(1–290)-affinity, DNA–cellulose and gel-filtration columns. When the proteins in this fraction were analysed by LC-MS/MS (liquid chromatography tandem MS), CK1δ/ε (casein kinase 1δ/ε) was the only protein kinase identified. Recombinant CK1δ/ε was found to bind to the N-terminal domain of Dnmt1 and significantly phosphorylated this domain, especially in the presence of DNA. Phosphorylation analyses using various truncation and point mutants of Dnmt1 revealed that the major priming site phosphorylated by CK1δ/ε was Ser146, and that subsequent phosphorylation at other sites may occur after phosphorylation of the priming site. When the DNA-binding activity of phosphorylated Dnmt1 was compared with that of the non-phosphorylated form, phosphorylation of Dnmt1 was found to decrease the affinity for DNA. These results suggest that CK1δ/ε binds to and phosphorylates the N-terminal domain of Dnmt1 and regulates Dnmt1 function by reducing the DNA-binding activity.
Skip Nav Destination
Article navigation
May 2010
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
April 14 2010
The DNA-binding activity of mouse DNA methyltransferase 1 is regulated by phosphorylation with casein kinase 1δ/ε
Yasunori Sugiyama;
Yasunori Sugiyama
*Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan
Search for other works by this author on:
Naoya Hatano;
Naoya Hatano
†Integrated Center for Mass Spectrometry, Graduate School of Medicine, Kobe University, Kobe 657-8501, Japan
Search for other works by this author on:
Noriyuki Sueyoshi;
Noriyuki Sueyoshi
*Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan
Search for other works by this author on:
Isao Suetake;
Isao Suetake
‡Laboratory of Epigenetics, Institute for Protein Research, Osaka University, Osaka 565-0871, Japan
Search for other works by this author on:
Shoji Tajima;
Shoji Tajima
‡Laboratory of Epigenetics, Institute for Protein Research, Osaka University, Osaka 565-0871, Japan
Search for other works by this author on:
Eiji Kinoshita;
Eiji Kinoshita
§Department of Functional Molecular Science, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima 734-8553, Japan
Search for other works by this author on:
Emiko Kinoshita-Kikuta;
Emiko Kinoshita-Kikuta
§Department of Functional Molecular Science, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima 734-8553, Japan
Search for other works by this author on:
Tohru Koike;
Tohru Koike
§Department of Functional Molecular Science, Graduate School of Biomedical Sciences, Hiroshima University, Hiroshima 734-8553, Japan
Search for other works by this author on:
Isamu Kameshita
Isamu Kameshita
1
*Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan
1To whom correspondence should be addressed (email kamesita@ag.kagawa-u.ac.jp).
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
December 07 2009
Revision Received:
February 24 2010
Accepted:
March 01 2010
Accepted Manuscript online:
March 01 2010
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2010 Biochemical Society
2010
Biochem J (2010) 427 (3): 489–497.
Article history
Received:
December 07 2009
Revision Received:
February 24 2010
Accepted:
March 01 2010
Accepted Manuscript online:
March 01 2010
Citation
Yasunori Sugiyama, Naoya Hatano, Noriyuki Sueyoshi, Isao Suetake, Shoji Tajima, Eiji Kinoshita, Emiko Kinoshita-Kikuta, Tohru Koike, Isamu Kameshita; The DNA-binding activity of mouse DNA methyltransferase 1 is regulated by phosphorylation with casein kinase 1δ/ε. Biochem J 1 May 2010; 427 (3): 489–497. doi: https://doi.org/10.1042/BJ20091856
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Captcha Validation Error. Please try again.