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. 2016 Mar 30;15(5):667–677. doi: 10.1080/15384101.2016.1147631

Figure 2.

Figure 2.

MiR-15b and CPEB1 control WEE1 expression. (A) Depletion of CPEB1 by siRNAs (Western blot). (B) Depletion of miR-15b by an antisense LNA (QPCR); (C) Influence of CPEB1 and/or miR-15b depletion on WEE1 3′UTR reporter luciferase activity transfected into HeLa cells along with inhibitors as indicated. Controls were miRNA (Applied Biosystems) and AllStars (Qiagen). Numbers indicate the ratio LucR/LucF for each sample. (D) Localization of WEE1 mRNA, CPEB1, and miR-15b in HeLa cells. HeLa cells were transfected with Cy5-labeled miR-15b, CPEB1-Cherry, FITC-labeled WEE1 3′UTR (wt or mutant as indicated). Arrows point to granules with co-localized WEE1 mRNA, CPEB1, and/or miR-15b. (E) CPEB1 and miR-15b control WEE1 protein expression. For gain-of-function assays, HeLa cells were transfected with a construct expressing CPEB1-Cherry (or Cherry as a control), or with miR-15b precursor (or a control miRNA); for loss-of-function assays, cells were transfected with the inhibitors described in A and B; WEE1 expression was monitored 48h later by Western blot. (F) Overexpression of miR-15b monitored by QPCR. (G) Overexpression of CPEB1 monitored by Western blot.