UTR RNA-induced apoptosis is mediated by double-stranded RNA and independent on the triphosphate moiety.

<p>(A) Double-stranded structures contribute to the UTR RNA-triggered apoptosis. PK-15 cells were left untreated (mock), or transfected with <i>in vitro</i> transcribed untreated (−), RNaseIII-treated (III), or RNaseI-treated (I) 5′UTR-Luc-3′UTR RNA (5′-Luc-3′) for 16 h prior to DNA laddering assay. (B) PK-15 cells were mock-transfected (mock), or transfected with <i>in vitro</i> transcribed luciferase RNA (Luc), or with calf intestinal phosphotase-treated (+) or untreated (−) 5′UTR-Luc-3′UTR RNA (5′-Luc-3′) for 16 h prior to DNA laddering assay. M: DNA size marker. (C) The degree of apoptosis correlates with the level of suppression of translation. PK-15 cells were transfected with the p2luc reporter plasmid (0.5 µg) alone, or in combination with 0.05 pmole of 5′UTR, 5′UTR-Luc-3′UTR (5′LUC3′), 3′UTR, or control firefly luciferase (Luc) RNA. The expression of <i>Renilla</i> luciferase (from p2luc plasmid) was quantified 7 h post-transfection. (D) Activation of PKR is responsible for the shutoff of protein synthesis. Total protein of mock-transfected PK-15 cells (lane 1) or of cells transfected with Luc RNA (lane 2) or 5′UTR-Luc-3′UTR RNA (lane 3) was resolved by SDS-PAGE. The presence of phosporylated eIF-2α (eIF-2α–Pi) was determined by Western blot analysis.