Fig. 3.

Mouse cytomegalovirus (MCMV) infection and type-I interferon (IFN) reciprocally regulate Clec2d promoter activity. a Schematic representation of the Clec2d upstream regulatory region showing selected consensus motifs; ATG, translational start site; +1, transcriptional start site (TSS); TBP, TATA binding protein; IRF, interferon regulatory factor; Sp, specificity protein. b Nucleotide sequence of the cluster of predicted IRF-binding sites (IRF cluster) within the first 200 bp of the Clec2d promoter. The upstream GAAA motifs in bold may enhance STAT1 (and thus ISGF3) recruitment to the IRF9 motif, in turn minimizing the requirement for the IRF3/7 motifs in response to type-I IFN [30]. c, d NIH3T3 cells were transfected with luciferase reporter plasmids containing varying sizes of the Clec2d promoter region as outlined in a and b. c Stable transfectants were infected with MCMV at a multiplicity of infection of 0.5 PFU/cell for 24 h. d Transient transfectants were treated with 10³ U/mL IFN-α₄ for 24 h the next day. Promoter activity was assayed by luciferase reporter assay relative to total protein and empty vector (pGL4.22) (b) or Renilla and empty vector (pGL3) (c). Significance was determined by a 2-tailed t test on log-transformed values (n = 3–7 experiments). * p < 0.05, ** p < 0.01, *** p < 0.001.