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. 2010 Dec 8;5(12):e14258. doi: 10.1371/journal.pone.0014258

Figure 3. The HCV replicon suppresses IPS-1/DDX3-mediated augmentation of IFN promoter activation.

Figure 3

(A,B) O cells with the HCV replicon fail to activate an IFN-β reporter in response to IPS-1/DDX3. O cells contain the full-length HCV replicon, and Oc cells do not [16]. O cells (A) or Oc cells (B) were transfected with IPS-1, DDX3 or p125luc reporter plasmids. At timed intervals (24 hrs), reporter activity was determined as in Fig. 2. (C,D) The HCV replicon suppresses IFN-promoter activation by polyU/UC. O cells and Oc cells expressing IPS-1 and DDX3 were stimulated with polyU/UC. At 48 hrs, reporter activity was determined as in panel A. (E) DDX3 is required for enhanced activation of IFN-beta promoter by O cell HCV 3′UTR. HCV 3′ UTR cDNA was amplified by RT-PCR from RNA extracted from O cells containing full-length HCV replicon. The HCV 3′ UTR RNA was synthesized in vitro using T7 RNA polymerase. DDX3 siRNA or control siRNA was transfected into HEK293 cells with the p125luc reporter. After 24 hrs, cells were transfected with HCV RNA, and incubated for 24 hrs. The IFN-beta promoter activation was assessed by luciferase reporter assay. One representative of at least three independent experiments, each performed in triplicate, is shown.