Using growth rate histograms as reporters of clathrin dynamics. (A) Kymographs are generated from the same BSC-1 cell before and during microaspiration, respectively. Elongated AP2 traces demonstrate longer CCS lifetimes under increased membrane tension. (B) Growth rate distributions are shown for seven different cells before and during aspiration. Change in CCS dynamics induced by micropipette aspiration can be observed in growth rate distributions. In the control experiments (cells before aspiration), CCSs spend more time in the formation phase (i.e., the distribution is inclined toward positive slopes). The asymmetry was abolished upon aspiration and plateau phases got relatively longer (Ncells = 7 and Ntraces = 38136). (C) Growth rate distributions in B are assembled in five bins to better delineate different phases of clathrin-coated vesicle formation (FD, fast dissolution; FF, fast formation; P, plateau; SD, slow dissolution; SF, slow formation). The bars show mean + standard deviation to illustrate dispersion between cells. P-values were obtained using the two-tailed t test. (D) 2D histograms of normalized intensity traces aligned at different time points (beginning, trace maximum, and end) and superposed as represented by the cartoons. In each alignment, the aspirated cells show a significantly widened distribution, demonstrating a preference for slopes lower in magnitude. Bins corresponding to multiples of 12 s are more populated, as they contain trace data from both 3- and 4-s frame rate acquisitions.
