Figure 8. IFI6 loss results in DNA replication stress and apoptosis induction in MEL-ST/NRASQ61K cells.
(A) MEL-ST cells expressing empty vector or NRASQ61K were analyzed by RT-qPCR. The expression of each gene in MEL-ST/NRASQ61K cells is shown relative to expression in empty vector control. (B) MEL-ST/NRASQ61K cells with the indicated shRNA were analyzed for DNA replication using the DNA fiber assay. The percentages of ongoing (ON), newly fired (NF), and terminated (Tr) DNA forks are shown. (C) Growth In Low Attachment (GILA) assay was performed using MEL-ST/NRASQ61K cells expressing either non-silencing (NS) or IFI6 shRNA. Cell death (%) was measured 48 hr after plating under the indicated conditions using the trypan blue exclusion assay and plotted. (D) MEL-ST/NRASQ61K cells expressing either NS or IFI6 shRNA were plated on poly-HEMA plates. Cell death (%) was measured 48 hr after plating under each condition using the trypan blue exclusion assay and plotted. (E) MEL-ST/NRASQ61K cells expressing either NS or IFI6 shRNA were plated on poly-HEMA plates. Apoptotic cell death (%) was measured 48 hr after plating under each condition by annexin V-FITC staining and plotted. (F) MEL-ST/NRASQ61K cells expressing either NS or IFI6 shRNA were plated on poly-HEMA plates. After 48 hr, the indicated proteins were analyzed by immunoblotting. ACTINB was used as a control. In all panels, data are presented as mean ± SEM, and *p<0.05.
Figure 8—figure supplement 1. DNA fiber assay results in MEL-ST/NRASQ61K cells.
Figure 8—figure supplement 2. IFI6 knockdown in MEL-ST/NRASQ61K does not induce senescence.
