Differentiation of T lymphocytes takes place in the thymus in a process that involves programmed activation of multiple genetic loci. Some of these genes encode constitutive products of mature T cells, such as components of their antigen-binding receptors. Other genes of interest are inducible genes that mediate T-cell responses after the cells have been triggered by antigen, such as those encoding interleukin 2 (IL2) and the IL2 receptor. The goal of this proposal is to study the sequence of activation of T-cell-specific genes in the thymus, with a particular focus on the second (inducible) class. The experiments will be designed to determine separately when the genes first become inducible at the chromatin level, and when the cells develop the signal-transduction mechanism that can turn these genes on in response to stimuli such as antigen recognition. The basis for the work is recent evidence that the IL2 gene itself becomes inducible before the develpoing T cell expresses receptors that can trigger induction. In situ hybridization will be used to detect IL2 induction in individual cells, and flow cytometry using the Ca2+-sensitive dye indo-1 will be used to measure their mobilization of intracellular Ca2+ in response to mitogenic lectins. Expression of high-affinity IL2 receptors will be measured by binding of radiolabeled IL2, adapted to autoradiography for single-cell detection. These methods will be used to dissect heterogeneity in the immature precursor thymocyte class, and to analyze the apparent loss of competence in cells becoming committed to the sterile, major-cortical differentiation pathway. Finally, preliminary results suggest that immature precursor thymocytes can be maintained in culture without destroying their abiltiy to differentiate. A vital aim of the proposal is therefore to optimize these culture conditions so that thymic lineage relationships can be determined and differentiation pathways manipulated directly, under controlled conditions in vitro.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI019752-07
Application #
3129154
Study Section
Immunobiology Study Section (IMB)
Project Start
1982-06-01
Project End
1989-08-31
Budget Start
1988-09-01
Budget End
1989-08-31
Support Year
7
Fiscal Year
1988
Total Cost
Indirect Cost
Name
California Institute of Technology
Department
Type
Schools of Arts and Sciences
DUNS #
078731668
City
Pasadena
State
CA
Country
United States
Zip Code
91125
Yang-Snyder, J A; Rothenberg, E V (1998) Spontaneous expression of interleukin-2 in vivo in specific tissues of young mice. Dev Immunol 5:223-45
Scherer, L J; Diamond, R A; Rothenberg, E V (1994) Developmental regulation of cAMP signaling pathways in thymocyte development. Thymus 23:231-57
Rothenberg, E V (1994) Signaling mechanisms in thymocyte selection. Curr Opin Immunol 6:257-65
Rothenberg, E V; Diamond, R A (1994) Costimulation by interleukin-1 of multiple activation responses in a developmentally restricted subset of immature thymocytes. Eur J Immunol 24:24-33
Rothenberg, E V; Diamond, R A; Chen, D (1994) Programming for recognition and programming for response. Separate developmental subroutines in the murine thymus. Thymus 22:215-44
Rothenberg, E V; Chen, D; Diamond, R A (1993) Functional and phenotypic analysis of thymocytes in SCID mice. Evidence for functional response transitions before and after the SCID arrest point. J Immunol 151:3530-46
Chen, D; Rothenberg, E V (1993) Molecular basis for developmental changes in interleukin-2 gene inducibility. Mol Cell Biol 13:228-37
Yang-Snyder, J A; Rothenberg, E V (1993) Developmental and anatomical patterns of IL-2 gene expression in vivo in the murine thymus. Dev Immunol 3:85-102
Rothenberg, E V (1992) The development of functionally responsive T cells. Adv Immunol 51:85-214
Novak, T J; Yoshimura, F K; Rothenberg, E V (1992) In vitro transfection of fresh thymocytes and T cells shows subset-specific expression of viral promoters. Mol Cell Biol 12:1515-27

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