Regulation of RAG-1 and CD69 expression in the thymus during positive and negative selection.
Successful interaction of the T cell receptor (TCR) with major histocompatibility complex (MHC) molecules during thymic selection down-regulates the expression of the recombination activating genes (RAG)-1 and -2 in cortical thymocytes and thereby prevents further endogenous TCR alpha-chain gene rearrangements (Borgulya, P., Kishi, H., Uematsu, Y. and von Boehmer, H., Cell. 1992. 69: 529-537; Brändle, D., Müller, C., Rülicke, T., Hengartner, H. and Pircher, H., Proc. Natl. Acad. Sci. USA 1992. 89: 9529-9533). To address the question whether down-regulation of RAG-1 activity represents an irreversible process we have blocked TCR-MHC interactions of thymocytes with thymic stromal cells. Firstly, transgenic (Tg) mice expressing a virus-specific MHC class I (H-2Db)-restricted TCR were injected with anti-Db or anti-CD8 monoclonal antibodies and RAG-1 expression was examined by in situ hybridization on thymus sections. The results show that cortical thymocytes up-regulated RAG-1 expression within 24 h after antibody administration. Secondly, immature thymocytes from TCR Tg mice were released from the thymic microenvironment and cultured in vitro for 14 h in single-cell suspension. The amount of RAG-1 mRNA was increased sixfold in cultured cells when compared to freshly isolated thymocytes. In addition, we show that immature thymocytes from TCR transgenic mice bearing non-selective MHC molecules (H-2d) down-regulated RAG-1 expression after antigen-induced TCR engagement. Cytofluorometric analysis further revealed that surface expression of CD69 on immature thymocytes inversely correlated with RAG-1 expression during positive and negative selection processes.
Brändle
D
D
Institute of Experimental Immunology, University of Zürich, Switzerland.
Müller
S
S
Müller
C
C
Hengartner
H
H
Pircher
H
H
D000818:Animals; D000911:Antibodies, Monoclonal; D015703:Antigens, CD / Q000096:biosynthesis*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000096:biosynthesis*; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000502:physiology; D005434:Flow Cytometry; D006183:H-2 Antigens; D018398:Homeodomain Proteins; D017403:In Situ Hybridization; D037181:Lectins, C-Type; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D014176:Protein Biosynthesis; D016693:Receptors, Antigen, T-Cell, alpha-beta; D013601:T-Lymphocytes / Q000378:metabolism* / Q000502:physiology; D013950:Thymus Gland / Q000166:cytology* / Q000378:metabolism*; D000099192:CD69 Antigens
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D006183:H-2 Antigens; D018398:Homeodomain Proteins; D037181:Lectins, C-Type; D016693:Receptors, Antigen, T-Cell, alpha-beta; D000099192:CD69 Antigens; C064658:RAG-1 protein