Rapid ANO5 accumulation at injury sites is not explained by FRAP or localized fiber contraction. (A) Time constants and associated errors from single exponential fits of ANO5 or dysferlin accumulation time courses (from Fig. 2). (B) Deconvolved images showing human ANO5-tomato after bleaching with an 8-s 561-nm laser pulse. (C) Quantification of human ANO1 fluorescence at wound-adjacent sarcolemma following 405-nm laser injury or ANO5 fluorescence after 561-nm laser bleaching in WT mouse myofibers. Fluorescence is normalized to prelaser fluorescence values. Time course of human ANO5 accumulation in WT fibers (from Fig. 2) is shown as a blue dotted line for comparison. ANO1 n = 10 fibers pooled from two mice, ANO5 n = 8 fibers from one mouse. (D) Images showing the contribution of localized myofiber contraction after injury to the apparent accumulation of plasma membrane proteins at the wound site. (E and F) Line profile analysis of ANO1 (E) or ANO5 (F) fluorescent intensity before (Initial) or after (Max) laser injury. Fluorescence values are measured from dotted lines as shown in top-row images of D. “Distance” is in units of micrometers along the analysis line, centered on the maximal relative fluorescence. ANO1 n = 8 fibers pooled from two mice, ANO5 n = 9 fibers pooled from four mice. Data are mean ± SEM. Scale bar = 10 μm.