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. 2020 Mar 13;295(16):5335–5349. doi: 10.1074/jbc.RA119.012260

Figure 4.

Figure 4.

PRR11 promotes F-actin polymerization via both N and C termini. a, subcellular distribution of truncated mutations. H1299 cells were overexpressed with Flag-tagged pvN173, WT PRR11, Δ33–41, Δ1–100, Δ100–200, Δ200–300, Δ290–360, Δ185–200, Δ100–184, or Δ1–100/200–360, and then the cells were fixed 24 h after and stained for Flag. Representative images are shown. Bar, 10 μm. Zoomed images of boxed region are shown at the bottom-left corner (scale bars, 5 μm). b, quantitative analysis of circular granules demonstrated in a. The number of circular granules per Flag-positive cells is given. Results are shown as the mean ± S.E. (n = 3). Greater than 20 cells were counted per condition in every repeat. c–e, effects of truncated mutations on F-actin polymerization and rearrangement. c, H1299 cells were overexpressed with Flag-tagged pvN173, WT PRR11, Δ1–100, Δ100–200, Δ290–360, Δ185–200, Δ100–184, or Δ1–100/200–360, and then cells were fixed 24 h after and stained for Flag and phalloidin. Representative images are shown. Bar, 10 μm. Zoomed images of boxed region are shown at the bottom-left corner (scale bars, 5 μm). d, quantitative analysis of aberrant F-actin polymerization cells demonstrated in c. Ratio indicated that the number of cells with aberrant F-actin polymerization is relative to the number of Flag-positive cells. Results are shown as the mean ± S.E. (n = 3). Greater than 20 cells were counted per condition in every repeat. e, quantitative analysis of actin comet tails demonstrated in c. The number of cells with actin comet tails per Flag-positive cells is shown. Results are shown as the mean ± S.E. (n = 3). Greater than 20 cells were counted per condition in every repeat. f, CK-666 inhibited actin comet tails but not circular granules induced by Δ100–200. H1299 cells were overexpressed with Δ100–200 and combined with/without CK-666 (84 μm), and then cells were fixed 24 h after and stained for Flag and phalloidin. Representative images are shown. Bar, 10 μm. Zoomed images of boxed region are shown at the bottom-left corner (scale bars, 5 μm). g, quantitative analysis of circular granules demonstrated in f. The number of circular granules per Flag-positive cells is shown. Results are shown as the mean ± S.E. (n = 3). Greater than 20 cells were counted per condition in every repeat. h, quantitative analysis of actin comet tails demonstrated in f. The number of cells with actin comet tails per Flag-positive cells is shown. Results are shown as the mean ± S.E. (n = 3). Greater than 20 cells were counted per condition in every repeat. Cell nuclei were stained with DAPI (blue).***, p < 0.001, or N.S., not significant (p > 0.05).