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. 2015 Feb 9;11(2):787. doi: 10.15252/msb.20145808

Figure 5. IFIX interactions with PML bodies and DNA damage effectors reflect antiviral properties.

Figure 5

  • A SAINT-filtered IFIX interactions related to PML body function or DNA damage response were assessed by STRING and visualized in Cytoscape. Node colors correspond to the log2-transformed NSAF/PAX values. Several interactions failing to pass SAINT, but enriched ≥ twofold relative to GFP control immunoisolations were added manually (gray color).
  • B Co-localization of IFIX-GFP with selected interactions in HEK293 cells was assessed by fluorescence confocal microscopy (white arrows). Scale bars, 5 μm.
  • C IFIX-GFP-expressing HEK293 cells were infected at MOI = 0.1 with HSV-1. At 72 h post-infection, progeny virus was harvested and titered. Mean values ± SEM (n = 3) are shown. *P ≤ 0.05, compared to control GFP cells.
  • D PML knockdown efficiency in HFFs relative to shScrambled (shSCR)-HFFs was assessed by Western blot (left). shPML-HFFs were infected at MOI = 0.1 with HSV-1. Progeny virus was harvested at 48 h post-infection and titered (right). Mean values ± SEM (n = 3) are shown. **P ≤ 0.01, compared to shSCR-HFFs.
  • E ifixmRNA levels in shIFIX-HFFs were assessed by RT–qPCR to determine knockdown efficiency (left). mRNA levels were normalized to cellular β-actin levels. The basal level of ifix expression in shSCR HFF cells relative to actin was 1.8E-5, which corresponds to a raw Ct value of ˜31. shIFIX-HFFs were infected at MOI = 0.1 with HSV-1. Progeny virus was harvested at 48 h post-infection and titered (right). Mean values ± SEM (n = 3) are shown. *P ≤ 0.05, compared to shSCR-HFFs.

Data information: In (C–E), Student's unpaired t-test; two-tailed.