WT and PM (S38D) Ana2 accumulate on procentrioles but the phospho-null Ana2-S38A mutant does not. (A–C) Interphase centrioles in transgenic GFP-Ana2–expressing cells were imaged using super-resolution microscopy. S2 cells were depleted of endogenous Ana2 for 12 d by transfection of UTR dsRNA on days 0, 4, and 8. On days 4 and 8, cells were cotransfected with the indicated GFP-Ana2 constructs (green) and induced to express on day 5 by addition of 0.1 mM CuSO₄. On day 12, cells were immunostained with anti-PLP (red) to mark the surface of mature centrioles. DNA was visualized with Hoechst (not depicted). Images are grouped as centrioles possessing two (A), one (B), or no (C) Ana2 spots. Bars, 200 nm. Right: Graphs of PLP (red) and GFP-Ana2 (green) fluorescence intensities plotted against the length of a linescan (white arrow) spanning the diameter of an individual centriole. (D) Graph of Ana2 fluorescence intensity as a ratio of procentriole to mother centriole spots in centrioles containing two spots. Asterisks mark significant differences between treatments: *, P < 0.05; **, P < 0.01. Error bars, SEM; n = 3 experiments, ∼5–12 centrioles per treatment.