ADAR1 Protein Induces Adenosine-targeted DNA Mutations in Senescent Bcl6 Gene-deficient Cells

Nobuhide Tsuruoka,Masafumi Arima,Nobuya Yoshida,Seiji Okada,Akemi Sakamoto,Masahiko Hatano,Hisae Satake,Eggi Arguni,Ji-Yang Wang,Jing-Hua Yang,Kazuko Nishikura,Souei Sekiya,Makio Shozu,Takeshi Tokuhisa

doi:10.1074/jbc.m112.365718

Abstract

Somatic mutations accumulate in senescent cells. Bcl6, which functions as a transcriptional repressor, has been identified as a potent inhibitor of cell senescence, but a role of Bcl6 in the accumulation of somatic mutations has remained unclear. Ig class-switch recombination simultaneously induces somatic mutations in an IgM class-switch (Ig-Sμ) region of IgG B cells. Surprisingly, mutations were detected in the Ig-Sμ region of Bcl6-deficient IgM B cells without class-switch recombination, and these mutations were mainly generated by conversion of adenosine to guanosine, suggesting a novel DNA mutator in the B cells. The ADAR1 (adenosine deaminase acting on RNA1) gene was overexpressed in Bcl6-deficient cells, and its promoter analysis revealed that ADAR1 is a molecular target of Bcl6. Exogenous ADAR1 induced adenosine-targeted DNA mutations in IgM B cells from ADAR1-transgenic mice and in wild-type mouse embryonic fibroblasts (MEFs). These mutations accumulated in senescent MEFs accompanied with endogenous ADAR1 expression, and the frequency in senescent Bcl6-deficient MEFs was higher than senescent wild-type MEFs. Thus, Bcl6 protects senescent cells from accumulation of adenosine-targeted DNA mutations induced by ADAR1.

Highlights

Bcl6 is a potent inhibitor of cell senescence, and somatic mutations accumulate in senescent cells
2 The abbreviations used are: mouse embryonic fibroblasts (MEFs), mouse embryonic fibroblast; Activation-induced cytidine deaminase (AID), activation-induced cytidine deaminase; GC, germinal center; Ig-S␮, IgM classswitch; ADAR1, adenosine deaminase acting on RNA1; Tg, transgenic; ADAR1-cKO, ADAR1-conditional deficient; P, passage; NP-CG, (4-hydroxy3-nitrophenyl)acetyl-chicken ␥ globulin; Somatic Hypermutation (SHM), somatic hypermutation; Ab, antibody; Quantitative Reverse Transcription (qRT), real-time quantitative RT
Somatic Mutations in the Ig-S␮ Region and the c-myc Gene of Bcl6-KO B Cells—Naive IgM B cells from spleens of WT and Bcl6-KO mice were stimulated with LPS and IL-4 for 4 days, and IgM B cells and IgG1 B cells were isolated by fluorescence activated cell sorter (FACS)

Summary

Background

Bcl is a potent inhibitor of cell senescence, and somatic mutations accumulate in senescent cells. ADAR1 overexpression induced adenosine-targeted DNA mutations in the Ig-S␮ region of IgM B cells from spleens of ADAR1 transgenic (Tg) mice and in the Ig-S␮ region and the c-myc gene of WT MEFs without induction of AID. These mutations accumulated in MEFs at a senescent stage accompanied with endogenous ADAR1 expression, and the frequencies in senescent Bcl6-KO MEFs were higher than senescent WT MEFs. We discuss a physiologic role of Bcl in protection of senescent cells against accumulation of adenosine-targeted DNA mutations induced by ADAR1

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION