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Nat Med. 2001 Oct;7(10):1155-8.

The analysis of costimulatory receptor signaling cascades in normal T lymphocytes using in vitro gene transfer and reporter gene analysis.

Author information

1
Department of Immunology, Mayo Clinic, Rochester, Minnesota, USA.

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.

PMID:
11590441
DOI:
10.1038/nm1001-1155
[Indexed for MEDLINE]

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