Fig. 6 | Nature Communications

Fig. 6

From: Membrane-cytoskeletal crosstalk mediated by myosin-I regulates adhesion turnover during phagocytosis

Fig. 6

Myo1e/f regulate actin dynamics at phagocytic adhesion sites. a Actin waves dynamics in WT and dKO macrophages transfected with EGFP-actin performing frustrated phagocytosis and imaged by TIRFM. Montage has been color-coded with respect to time (minutes:seconds) to display adhesion turnover. The white line marks the cell’s edge. The final images at the right (Merge) are maximum intensity projections. Scale bars, 2 μm. See also Supplementary Movie 13. b Gray scale zoom of maximum intensity projections in a to show distinct punctate structure of WT adhesions compared to fused/enlarged adhesions in dKO macrophage. Scale bar, 2 μm. c Graph of FRAP curves (mean ± SD). WT and dKO BMDM transfected with EGFP-actin were subjected to FRAP analysis to measure actin turnover during frustrated phagocytosis. The resulting data was corrected for photobleaching, normalized to the pre-bleached images, then fit using a single exponential function. Data pooled from three independent experiments (n = 38 WT and 36 dKO cells). d Representative actin wave recovery assessed by FRAP. White box indicates bleached region. Scale bar, 1 μm. e Graph of half-life of recovery (mean ± SD). Data from three independent experiments, (p = 0.39, unpaired t-test). f Graph of mobile fraction (mean ± SD). Data from three independent experiments, p = 0.0035, unpaired t-test). g Time-lapse montage of tracking analysis to characterize dKO macrophage actin wave dynamics. The border of the actin wave was tracked for protrusion (green)/retraction (yellow) speed and the inner blue line marks the inner boundary for actin quantification. Time, minutes:seconds. Scale bar, 5 μm. See also Supplementary Movie 14. h Increased actin wave intensity correlated with stalled actin wave. Graph (mean ± SD) showing the distribution of intensities of actin adhesions plotted against actin wave boundary speed during frustrated phagocytosis in a dKO BMDM. Error bars represent variability between individual boundary speeds measurements. See also Supplementary Figure 10

Back to article page