Figure S3.
The development of periodic actomyosin-II impacts the capacity of stress-induced reversible axon beading. (A) Representative SIM images illustrating p-MRLC periodicity measurement (see Materials and methods). The traced lines of p-MRLC and the non-periodic β III tubulin are shown in the bottom panels. Bar = 1 µm. (B) The line profiles of traced p-MRLC (blue line) and β III tubulin (black line) from A. p-MRLC and β III tubulin peaks are indicated with red circles, respectively. (C) The ratio of periodic p-MRLC is calculated as peaks with spacing between 0.2 ± 0.025 µm. (D) SIM images showing p-MRLC ring distribution in growth cones and distal axon shafts at DIV4 and DIV8 hippocampal neurons in AoC. Red brackets indicate gaps lacking periodic p-MRLC. Bar = 5 µm (top), 1 µm (bottom). (E) Comparison of periodicity ratio between growth cones and distal axon shafts in D (N = 18, 23, 21, 27). (F) Time-lapse images showing the beading process in growth cones (GC, 1#) and axon shafts (2#) before and after flux at DIV4 and DIV8. Bracketed ROIs amplified in bottom panels. Bar = 20 µm (top), 5 µm (bottom). (G) Paired comparison of flux-induced beading density as shown in F (N = 9, 11). (H) Positive correlation between p-MRLC periodicity and stress-induced axon beading density in growth cones or shafts on DIV4 or DIV8, with Pearson’s correlation coefficient close to 1. (I and J) Quantification of (I) initial bead number and (J) peak bead number during stress induced by 50 µl/min flux, related to Fig. 5 M. (N = 21, 41, 31, 42 in I; 21, 40, 32, 42 in J). Data represent mean ± SEM; in E, Tukey’s multiple comparisons test; in G, within groups paired two-tailed Student’s t test, between groups unpaired two-tailed Student’s t test; in I and J, Mann-Whitney test; *P < 0.05, **P < 0.01, ***P < 0.001.

The development of periodic actomyosin-II impacts the capacity of stress-induced reversible axon beading. (A) Representative SIM images illustrating p-MRLC periodicity measurement (see Materials and methods). The traced lines of p-MRLC and the non-periodic β III tubulin are shown in the bottom panels. Bar = 1 µm. (B) The line profiles of traced p-MRLC (blue line) and β III tubulin (black line) from A. p-MRLC and β III tubulin peaks are indicated with red circles, respectively. (C) The ratio of periodic p-MRLC is calculated as peaks with spacing between 0.2 ± 0.025 µm. (D) SIM images showing p-MRLC ring distribution in growth cones and distal axon shafts at DIV4 and DIV8 hippocampal neurons in AoC. Red brackets indicate gaps lacking periodic p-MRLC. Bar = 5 µm (top), 1 µm (bottom). (E) Comparison of periodicity ratio between growth cones and distal axon shafts in D (N = 18, 23, 21, 27). (F) Time-lapse images showing the beading process in growth cones (GC, 1#) and axon shafts (2#) before and after flux at DIV4 and DIV8. Bracketed ROIs amplified in bottom panels. Bar = 20 µm (top), 5 µm (bottom). (G) Paired comparison of flux-induced beading density as shown in F (N = 9, 11). (H) Positive correlation between p-MRLC periodicity and stress-induced axon beading density in growth cones or shafts on DIV4 or DIV8, with Pearson’s correlation coefficient close to 1. (I and J) Quantification of (I) initial bead number and (J) peak bead number during stress induced by 50 µl/min flux, related to Fig. 5 M. (N = 21, 41, 31, 42 in I; 21, 40, 32, 42 in J). Data represent mean ± SEM; in E, Tukey’s multiple comparisons test; in G, within groups paired two-tailed Student’s t test, between groups unpaired two-tailed Student’s t test; in I and J, Mann-Whitney test; *P < 0.05, **P < 0.01, ***P < 0.001.

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