Fig. 5.

Cyclic dinucleotides have no toxic effect on S2 cells, and are not likely to be second messengers in the cell death response to DNA. a Electroporation of S2 cells with CT DNA and cyclic dinucleotides. Cells were electroporated with 10 µg of CT DNA or 20 µg of cyclic di-GMP or cyclic di-AMP, and MTT cleavage was measured 1 h after treatment. Results show the mean and range of duplicate electroporations. b Transfection of S2 cells with CT DNA and cyclic dinucleotides using Lipofectamine 2000; 2 µg of CT DNA and 4 µg of cyclic dinucleotides were transfected into S2 cells complexed with 4 µl of Lipofectamine 2000, and MTT cleavage was measured 18 h after transfection. The bars show data from 4 experiments (mean ± SEM). c Induction of IFN-β mRNA in BMM either untreated (no ZAP) or 2 h after electroporation with either no addition, CT DNA or various amounts of cyclic di-GMP. The real time PCR results shown are mean and range of duplicate assays of IFN-β mRNA relative to HPRT.