Figure 4.

IFT20 moves in cell bodies and cilia. (A and B) Time-lapse fluorescence images of cells LLC-PK1 cells expressing IFT20-GFP were taken every 0.75 s for 26 s. These were adjusted for contrast and assembled into a movie that plays at ∼5× normal speed. (A) To illustrate the two types of movements observed, three frames (12–14) are shown. The longest arrow marks the basal body. The broad arrow marks IFT20 in the Golgi complex. The three small arrows mark the position of a particle moving from the basal body to the ciliary tip (anterograde movement). The position of the particle in each frame is marked by an asterisk (*). The arrowhead marks the position of a focus of IFT20-GFP that is moving rapidly in the cytoplasm. In the movie, this focus appears near the basal body, moves out into the cell, and then returns back to the basal body. Size bar, 2 μm long. (B) Kymographs showing IFT20-GFP movement in the two cilia in the movie. The top panel of each pair shows the raw kymographs, whereas lines have been drawn on the bottom panel to mark the moving particles. Anterograde particles show upward sloping lines, whereas retrograde particles show downward sloping lines. (C) Five image Z-stacks were taken of IMCD3 cells expressing IFT20-GFP every 150 ms. The Z-stacks were deconvolved, reduced to a single plane by summing, and assembled into a movie. In C three sequential frames from the movie were pseudocolored red, green, or blue (images 2, 3, and 4), and the three images were combined (images 2–4). The discrete colors in the combined image indicate that the IFT20-GFP is highly dynamic. The arrow in image 3 points to a tubular structure moving into the cilium.