Ciliary trafficking of Gli2 in the absence of Sufu function.

<p>Ciliary localization of endogenous Gli2 is undetectable in <i>Sufu</i>^<i>-/-</i> MEFs even though Gli2 shuttles in and out of cilia. (A) Immunofluorescence staining of <i>Sufu</i>^<i>-/-</i> MEFs and <i>Sufu</i>^<i>-/-</i> MEFs rescued with human SUFU (hSUFU) for Gli2 (green) and acetylated Tubulin (AcTub, red) and DAPI (blue). (B) Immunofluorescence staining of <i>Sufu</i>^<i>-/-</i> MEFs stably transfected for expression of Dync2hc or control shRNA with antibodies against Gli2 (green) and AcTub (red). (C) Unlike Gli2, a block of retrograde transport in primary cilia by <i>Dync2hc</i> shRNA does not increase ciliary accumulation of Sufu. NIH3T3 cells stably expressing HA-Gli2 with shRNA targeting either Dync2h1 or control were stained with antibodies against AcTub (green) and HA or Sufu (red). Arrowheads denote ciliary tip localization. A representative experiment from at least three independent experiments is shown. Ciliary tip localization of Gli2, Sufu, and HA-Gli2 was scored from more than 50 cells per each replicate and plotted in graphs right to immunofluorescence images. Error bars show mean +/- standard deviation. Statistical significance was measured by Student’s <i>t</i>-test: **** (P<0.0001), *** (0.00010.05).