Abstract
Ligation of the antigen receptor and costimulatory receptors on the surface of T lymphocytes initiates intracellular signals that regulate cell-cycle progression and cell differentiation. To effectively manipulate the activation of T cells for immunotherapeutic applications, it will be important to understand how these signaling pathways are integrated to control specific gene transcription events. Here we describe a novel transient transfection procedure that efficiently introduces DNA into non-dividing normal human and murine T lymphocytes while maintaining high cell viability. Using this technique, reporter genes can be introduced to characterize intracellular signaling pathways that regulate specific gene transcription events in normal T-lymphocyte populations. We show that the CD28 receptor can be differentially coupled to downstream signaling pathways in different T-lymphocyte populations. In addition, we demonstrate that a gene encoding a tagged constitutively active mitogen-activated kinase kinase-1 protein can be transfected and rapidly expressed to regulate the expression of Bcl-2 in normal thymocytes.
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This work was supported by funding from the American Cancer Society and the Mayo Foundation.
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Bell, M., Huntoon, C., Graham, D. et al. The analysis of costimulatory receptor signaling cascades in normal T lymphocytes using in vitro gene transfer and reporter gene analysis. Nat Med 7, 1155–1158 (2001). https://doi.org/10.1038/nm1001-1155
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DOI: https://doi.org/10.1038/nm1001-1155
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