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. 2016 Dec 30;18(2):241–263. doi: 10.15252/embr.201642386

Figure 2. Nup358 is required for miRNA function.

Figure 2

  1. Western analysis of HeLa cells, treated with control (siControl), Dicer (siDicer), Nup214 (siNup214), or Nup358 (siNup358) siRNA, for assessing the extent of protein depletion using indicated antibodies. Vinculin was used as loading control.
  2. HeLa cells were initially transfected with indicated siRNAs, followed by Renilla luciferase (RL) reporter constructs: RL‐control (no let‐7a binding site in the 3′‐UTR) or RL‐3xBulge (3 imperfect let‐7a binding sites in the 3′‐UTR). Firefly luciferase (FL) was co‐transfected to serve as internal control. RL/FL luminescence ratio was calculated. Data are presented as mean ± SD (n = 3), P‐values were calculated using Student's t‐test.
  3. HEK293T cells were transfected with indicated siRNAs, followed by pCMV‐FL‐miR30 (P) reporter with either pSUPER‐control or pSUPER‐miR30 constructs. RL was co‐transfected as internal control. FL/RL luminescence ratio was calculated. Data are presented as mean ± SD (n = 3), P‐values were calculated using Student's t‐test.
  4. HeLa cells were initially transfected with control (siControl), Dicer (siDicer), or Nup358 (siNup358)‐specific siRNAs, followed by FL constructs containing wild‐type (HMGA2‐wt) or mutant (HMGA2‐mut) HMGA2 3′‐UTR, along with RL as internal control. Graph was plotted using data from three independent experiments. FL/RL luminescence ratio was calculated. Data are presented as mean ± SD (n = 3), P‐values were calculated using Student's t‐test.