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. 2020 Sep 18;39(21):e104958. doi: 10.15252/embj.2020104958

Figure 3. Antisense oligonucleotides against the GA‐rich region specifically interfere with localization of Rab13 RNA .

Figure 3

  1. Schematic showing positions along the mouse Rab13 3′UTR targeted by the indicated PMOs. PMOs #1, 2, and 3 target the RGAAGRR motifs or the adjacent GA‐rich region. PMOs #6 and #7 target the Rab13 3′UTR outside of the GA‐rich region. The control PMO targets an intronic sequence of human β‐globin. Red rectangles and text indicate the location of the GA‐rich motifs.
  2. FISH images and corresponding PDI measurements of mouse fibroblast cells treated with the indicated PMOs. Cyb5r3 is an APC‐dependent RNA also enriched at protrusions. Arrows: peripheral Rab13 RNA. Arrowheads: perinuclear Rab13 RNA. Boxed regions are magnified in the insets. Note that Rab13 RNA becomes perinuclear in cells treated with PMOs against the GA‐rich region. Scale bars: 10 μm. (4 μm in insets). ****P < 0.0001 by analysis of variance with Dunnett's multiple comparisons test. N = 40–90 cells observed in 3–6 independent experiments. Bars: mean ± 95% CI.
  3. Protrusion (Ps) and cell body (CB) fractions were isolated from cells treated with control PMO or Rab13‐PMO #2. The indicated RNAs were detected through nanoString analysis to calculate Ps/CB enrichment ratios (n = 3; bars: mean ± s.e.m.). Note that only the distribution of Rab13 RNA is affected. **P = 0.01 by two‐way ANOVA with Bonferroni's multiple comparisons test against the corresponding control.
  4. Levels of the indicated RNAs were determined using nanoString analysis from control‐ or Rab13 PMO #2-treated cells (n = 4; bars: mean ± s.e.m.). No significant differences were detected by two‐way ANOVA against the corresponding controls.