Fig. 4: PALM localization and diffusion revealed by single-particle tracking.
a, Membrane localization of mEos3.2-Prli42 acquired from 2000 frames (left) and 5000 frames (right). b, and c, Localization of mEos3.2-RsbR1 in wild-type and Δprli42 strains, respectively (5000 frames). d, and e, Localization of mEos3.2-RsbL in wild-type and Δprli42 strains, respectively (5000 frames). The top and middle panels are bright-field and reconstructed images; Scale bars are 500 nm. Triangle symbols indicate high-density clusters. The cross-section profiles are shown in the lower panels. f, Distribution of single-molecule displacements of mEos3.2-Prli42. Blue and orange curves show the mBd and Cl fractions. The green dashed line is the sum of the two fractions. g, The DL of mEos3.2-Prli42 (N = 18), mEos3.2-RsbR1 (N = 26 and 10 for the wild-type and Δprli42, respectively) and mEos3.2-RsbL (N = 8 for both wild-type and Δprli42) were from cells in different fields of view (FOV) in three replicates. mEos3.2-Prli42 and mEos3.2-RsbR1 have membrane-bound (mBd) and clustered (Cl) fractions as shown on the x-axis; the starred membrane-bound fraction (mBd*) indicates mEos3.2-RsbR1 molecules loosely associated with membrane in the Δprli42 strain. mEos3.2-RsbL have free (Fr) and membrane-bound (mBd) fractions. Each dot shows the data of a FOV containing 10–50 well-separated cells. The box range indicates the standard deviation (SD), and the open square and dash symbols inside the boxes indicate the mean and median, respectively. h The corresponding percentage of the fraction in (g). Error bars represent standard deviations. Additional data on mEos3.2-RsbR1 and mEos3.2-RsbL localization and diffusion are in Supplementary Fig. 10.
