Skip to main content
. 2021 Apr 17;296:100680. doi: 10.1016/j.jbc.2021.100680

Figure 4.

Figure 4

Trafficking of post-Golgi and endosomal vesicles is increased upon targeting DIAPH3 to the base of cilia.A–G, targeting DIAPH3 to the base of cilia increased cargo trafficking. hTERT-RPE1 cells were transfected with GFP-CNTN1 or GFP-CNTN1-DIAPH3, and after 24 h of serum starvation, cells were fixed and stained for acetylated tubulin, GFP and (A) Arf4, (B) IFT20, (C) Rab11, (D) BBS1, or (E) GFP. Scale bars are 3 μm for each image. F, quantifications of the total protein fluorescence intensity scan at the base of cilia was conducted for Arf4, IFT20, Rab11, BBS1, or GFP. G, DIAPH3 targeted to the ciliary base via PACT increases the recycling endosome (Rab11) protein level at the base of the cilia. Cells were transfected with GFP-PACT or GFP-PACT-DIAPH3 and after 24-h serum starvation, were fixed and stained for acetylated tubulin, GFP, and Rab11. Quantification of the Rab11 fluorescence protein intensity scan was conducted. For panels F and G, box and whisker plots (as defined in Fig. 1C) are shown. H, targeting DIAPH3 to the base of cilia has no effect on protein levels of Arf4 or IFT20. Cells were transfected with GFP-CNTN1 (control) or GFP-CNTN1-DIAPH3, and samples were western blotted for GFP, GAPDH, Arf4, and IF20. Three independent experiments were conducted with n = 50 cells each, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.005, ∗∗∗∗p < 0.001. Two-tailed t-test analysis was done to compare GFP-CNTN1 control with GFP-CNTN1-DIAPH3.