Fig. 1.
A quantitative model of macropinosome formation. (A) Typical phase-contrast images of macropinosome formation (white arrows). Irregular ruffles were phase-dense bands at the periphery of the cells (+0 sec to +40 sec), which changed into curved ruffles (+60 sec). A curved ruffle (white arrow) closed into a circular ruffle (+120 sec), then moved centripetally after a 100-second pause (+220 sec). These morphological transitions were divided into four stages of macropinosome formation: irregular, curved, circular and motile. Scale bar: 3 μm. (B) The size distribution of 65 macropinosomes, defined as the diameter at the first frame of the circular ruffle stage. (C) The frequency of macropinosome formation. The x-axis indicates the time of the beginning of the curved stage, relative to the addition of M-CSF. (D) Macropinosomes in the size range of 1.2- to 2.2-μm diameter were analyzed (n=41), with the beginning of the curved stage set to 0 seconds. (E) The subset of macropinosomes used to evaluate signaling (1.2-2.2 μm, 60-second curved-ruffle stage). The beginning of the curved-ruffle stage was set as time=0. (F) Diagram of the two stages of macropinosome closure. The top row shows a side view and the bottom row shows a top view (as observed in the microscope, A) of plasma membrane and macropinosome membranes. Ruffle closure is the formation of a complete circular ruffle, comprised entirely of plasma membrane; ruffle closure marks the beginning of the circular stage. Cup closure marks the separation of the macropinosome from the plasma membrane sometime after ruffle closure.