The purpose of this research proposal is to analyze the molecular mechanisms of somatic DNA recombination in lymphocytes. The recombination occurs in the gene system for the antigen-recognizing molecules of differentiating lymphocytes.
Specific aims of this proposed project includes: 1) To further purify an endonuclease which specifically cleaves Ig DNA segments around the recombination site. 2) To isolate and characterize DNA binding proteins which recognize two consesus sequences at the recombination site. 3) To investigate a hypothetical RNA molecule which might hold two Ig gene segments together during recombination. 4) To establish an in vitro recombination system for V-(D)-J joining. 5) To establish a retroviral vector system to analyze the Ig gene recombination. 6) To examine the Ig heavy-chain and T-cell receptor B-chain genes for the formation of non-germline elements, which occurs at the joining sites of V, D and J segments. In addition to standard molecular cloning and DNA sequencing techniques, we will make use of the recombinant retroviral vector system for introducing defined Ig and T-cell receptor gene segments to various cell types of the lymphocyte lineages. It is hoped that the proposed research will contribute to our understanding of lymphocyte differentiation in molecular terms, but also to the clerification of complex recombination processes required for the activation of antibody genes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI018790-04
Application #
3128209
Study Section
Immunobiology Study Section (IMB)
Project Start
1982-08-01
Project End
1988-08-31
Budget Start
1985-09-01
Budget End
1986-08-31
Support Year
4
Fiscal Year
1985
Total Cost
Indirect Cost
Name
University of California Berkeley
Department
Type
Schools of Arts and Sciences
DUNS #
094878337
City
Berkeley
State
CA
Country
United States
Zip Code
94704
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Davis, D D; Yoshida, K; Kingsbury, L et al. (1991) Circular DNA resulting from recombination between V-(D)-J joining signals and switch repetitive sequences in mouse thymocytes. J Exp Med 173:743-6
Matsuoka, M; Nagawa, F; Okazaki, K et al. (1991) Detection of somatic DNA recombination in the transgenic mouse brain. Science 254:81-6
Shirakata, M; Huppi, K; Usuda, S et al. (1991) HMG1-related DNA-binding protein isolated with V-(D)-J recombination signal probes. Mol Cell Biol 11:4528-36
Yoshida, K; Matsuoka, M; Usuda, S et al. (1990) Immunoglobulin switch circular DNA in the mouse infected with Nippostrongylus brasiliensis: evidence for successive class switching from mu to epsilon via gamma 1. Proc Natl Acad Sci U S A 87:7829-33
Matsuoka, M; Yoshida, K; Maeda, T et al. (1990) Switch circular DNA formed in cytokine-treated mouse splenocytes: evidence for intramolecular DNA deletion in immunoglobulin class switching. Cell 62:135-42
Okazaki, K; Sakano, H (1988) Thymocyte circular DNA excised from T cell receptor alpha-delta gene complex. EMBO J 7:1669-74
Matsuoka, M; Hagiya, M; Hattori, T et al. (1988) Gene rearrangements of T cell receptor beta and gamma chains in HTLV-I infected primary neoplastic T cells. Leukemia 2:84-90
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Okazaki, K; Davis, D D; Sakano, H (1987) T cell receptor beta gene sequences in the circular DNA of thymocyte nuclei: direct evidence for intramolecular DNA deletion in V-D-J joining. Cell 49:477-85

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