Fig. 6. Frequent actin nucleation by mDia1 around sites of vigorous actin disassembly. (A) Images of EGFP-mDia1F2 (left) and mPlum-AIP1 (middle) were acquired in a live cell. Cells were then fixed, and stained with phalloidin in order to visualize F-actin (right). Note the clusters of mDia1F2 speckles (arrowheads) around the area labeled strongly with AIP1, a cofactor of cofilin. (B,C) Frequency of mDia1 speckle appearance is biased toward the region of high AIP1 concentration. The images show the position of mDia1F2 appearance (B, diamonds) or mDia1Full speckle appearance (C, white dots), mDia1Full trajectories (C, pink circles) and the intensity of mPlum-AIP1 fluorescence (pseudocolor). In the graphs, the x, y position of individual pixels within an mPlum-AIP1 image was binned into ten equal areas according to their fluorescence intensity values. Intensity-sorted positions were then used to correlate the appearance of mDia1 speckles with the local intensity of mPlum-AIP1 fluorescence (B, n=158, 1 cell; C, n=405, 11 cells). mDia1F2 speckles frequently emerged in areas strongly labeled with mPlum-AIP1. Frequency of mDia1Full speckle appearance shows positive correlation with the intensity of mPlum-AIP1 (P<0.01). Scale bars: 5 µm.