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Molecular and Cellular Biology, May 2004, p. 3692-3702, Vol. 24, No. 9
0270-7306/04/$08.00+0     DOI: 10.1128/MCB.24.9.3692-3702.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

In Vitro Processing of the 3'-Overhanging DNA in the Postcleavage Complex Involved in V(D)J Joining

Tadashi Nishihara, Fumikiyo Nagawa, Hirofumi Nishizumi, Masami Kodama,{dagger} Satoshi Hirose, Reiko Hayashi, and Hitoshi Sakano*

Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, Bunkyo-ku, Tokyo 113-0032, Japan

Received 27 October 2003/ Returned for modification 4 December 2003/ Accepted 2 February 2004

The postcleavage complex involved in V(D)J joining is known to possess a transpositional strand transfer activity, whose physiological role is yet to be clarified. Here we report that RAG1 and RAG2 proteins in the signal end (SE) complex cleave the 3'-overhanging structure of the synthetic coding-end (CE) DNA in two successive steps in vitro. The 3'-overhanging structure is attacked by the SE complex imprecisely, near the double-stranded/single-stranded (ds/ss) junction, and transferred to the SE. The transferred overhang is then resolved and cleaved precisely at the ds/ss junction, generating either the linear or the circular cleavage products. Thus, the blunt-end structure is restored for the SE and variably processed ends are generated for the synthetic CE. This 3'-processing activity is observed not only with the core RAG2 but also with the full-length protein.

* Corresponding author. Mailing address: Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan. Phone: 81-3-5689-7239. Fax: 81-3-5689-7240. E-mail: sakano{at}mail.ecc.u-tokyo.ac.jp.

{dagger} Present address: Radiobiology Division, National Cancer Center Research Institute, Tsukiji, Chuo-ku, Tokyo 104-0045, Japan.

Molecular and Cellular Biology, May 2004, p. 3692-3702, Vol. 24, No. 9
0022-538X/04/$08.00+0     DOI: 10.1128/MCB.24.9.3692-3702.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

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  • Nagawa, F., Hirose, S., Nishizumi, H., Nishihara, T., Sakano, H. (2004). Joining Mutants of RAG1 and RAG2 that Demonstrate Impaired Interactions with the Coding-end DNA. J. Biol. Chem. 279: 38360-38368 [Abstract] [Full Text]  


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