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. 2022 Feb 24;23(4):e52775. doi: 10.15252/embr.202152775

Figure EV1. Ccdc108 is essential for multiciliogenesis in epidermis of the Xenopus embryo (related to Fig 1).

Figure EV1

  1. In vivo validation of morpholino efficiency. ccdc108 morpholino inhibits the expression of the GFP reporter containing the MO target site of ccdc108 at the 5’UTR.
  2. Scanning electron microscope images and graphical plot show slightly shorter cilia in Ccdc108‐depleted MCCs. Experiment was performed once and 50 cilia from 10 MCCs in different microscopic fields were scored. Mean ± s.d. values are presented.
  3. Representative confocal images and graphical plot display reduced ciliary Ac‐tub levels in ccdc108 CRISPR mutants. A total of 20 images of 20 embryos for each condition. Cell membranes (mGFP, purple) and cilia (Ac‐tub, green) were labeled with indicated antibodies. Quantitative data from three independent experiments were scored. Unpaired two‐tailed t‐test was performed (***P < 0.001). Mean ± s.d. values are also presented.
  4. Graphical plot and cartoons show that epidermal MCC cilia beat frequency (CBF) and cilia beat pattern (CBP) are both affected in ccdc108 morpholino‐treated Xenopus embryos from stage 27. The cartoons were generated based on the results of imaging live MCCs by high‐speed video microscopy (Movies [Link], [Link], [Link]). Two biologically independent experiments were performed. Greater than 15 MCCs from four embryos for each condition.
  5. Transverse section views show that Ccdc108 is dispensable for primary ciliogenesis in the Xenopus neural tube. Embryos at stage 30 were fixed, and stained with the acetylated tubulin antibody (red) and DAPI (blue). mRNA of a membrane‐bound form of GFP (mGFP; green) was co‐injected with each morpholino to indicate targeted cells. Arrows mark primary cilia. Three biologically independent experiments were performed and images from the same experiment were presented.
  6. Effects of Ccdc108 depletion on motile monocilia formation and length in the gastrocoel roof plate (GRP). Embryos were fixed and stained with the acetylated tubulin antibody (red). mGFP (green) was co‐injected with each morpholino to indicate targeted cells. GRP explants were prepared from embryos at stage 18. Greater than 6 embryos from three biologically independent experiments for each condition were images and 10 cilia from each embryo were scored. Unpaired two‐tailed t‐test was performed. Mean ± s.d. values are also presented.

Source data are available online for this figure.