Research Paper Volume 5, Issue 5 pp 347—356

Figure 3. FOXO1 induction of miRNA expression depends on RNA polymerase II, but is independent of de novo protein synthesis. (A-B) Induction of miR-506 (A) and miR-507 (B) is prevented by an inhibitor of RNA polymerase II. HEK-293T cells expressing The cells were treated overnight as indicated. For combined treatment, α-amanitin was added 2 hours prior to addition of 4-hydoxitamoxifen (4HT). The levels of miR-506 (A) and miR-507 (B) were compared by quantitative PCR using RNU6B as an internal control and are shown relative to those in untreated cells. (C) Activation of FOXO1 induces miR-506 expression without de novo protein synthesis. HEK-293T cells harboring FOXO1-AAA-ER or the corresponding empty vector were treated for 8 hours with 4-hydoxitamoxifen (4HT), cycloheximide (protein synthesis inhibitor, CHX) or a combination thereof. The levels of miR-506 in treated and untreated cells were compared by quantitative PCR using RNU6B as an internal control and are shown relative to those of untreated vector-infected cells. An apparent increase in miR-506 levels in cycloheximide-treated vs. -untreated samples is due to a small decrease in RNU6B expression in the presence of cycloheximide (data not shown).