Fig. 5.

Tracking diffusion of IRE1 clusters. Clusters from a live-imaging experiment (A) with a 1-Hz frame rate are tracked to yield single-cluster trajectories (B); colors represent individual cluster trajectories. (Scale bars, 10 μm.) The trajectory of each cluster is broken up into individual 9-s chunks (C), which are then averaged to provide a separate MSD plot for each cluster (D). The MSD plot is fit with a straight line to yield a per-cluster diffusion constant (red line). Values shown are mean and 95% CI. (E) Log–log plot of calculated diffusion constants for each cluster plotted against the integrated fluorescence intensity of that cluster, which we take to be directly proportional to the number of IRE1 molecules in that cluster or the square root of the cluster’s radius. The best-fit linear regression line (blue) shows a minimal correlation between cluster intensity and diffusion constant, while enforcing a 1/R (orange) or 1/R² (green) dependence results in substantially worse fits. (F) Residual plots of the 3 fits in E. A.U., arbitrary units.