An adenoviral vector for probing promoter activity in primary immune cells

作者：Tripathi P; Madan R; Chougnet C; Divanovic S; Ma XJ; Wahl LM; Gajewski T; Karp CL; Hildeman DA^*

来源：Journal of Immunological Methods, 2006, 311(1-2): 19-30.

DOI：10.1016/j.jim.2006.01.009

• T cell
• macrophage
• dendritic cell
• transcription
• promoter

Functional analysis of the DNA regulatory regions that control gene expression has largely been performed through transient transfection of promoter-reporter constructs into transformed cells. However, transformed cells are often poor models of primary cells. To directly analyze DNA regulatory regions in primary cells, we generated a novel adenoviral luciferase reporter-vector, pShuttle-luciferase-GFP (pSLUG) that contains a promoterless luciferase cassette (with an upstream cloning site) for probing promoter activity, and a GFP expression cassette that allows for the identification of transduced cells. Recombinant adenoviruses generated front this vector can transduce a wide range of primary immune cells with high efficiency, including human macrophages. dendritic cells and T cells; and mouse T cells transgenic for the coxsackie and adenoviral receptor (CAR). In primary T cells, we show inducible nuclear factor of activated T cells (NF-AT) activity using a recombinant pSLUG adenovirus containing a consensus NF-AT promoter. We further show inducible IL-12/23 p40 promoter activity in primary macrophages and dendritic cells using a recombinant pSLUG adenovirus containing the proximal human IL-12/23 p40 promoter. The pSLUG system promises to be a powerful tool for the analysis of DNA regulatory regions in diverse types of primary immune cells.

• 出版日期2006-4-20