Figure 5.

Centriole Numbers Are Preserved in JS (V194A) and JATD (A199P) Mutant RPE-1 Cells

(A) CEP152 signal intensity and distribution are comparable among control, V194A, and A199P mutant RPE-1 cell lines. Cells stained by antibodies against the proximal centriole marker CEP152 and the nascent centriole marker HsSAS-6. Images correspond to maximum intensity projections of confocal micrographs of control, V194A, and A199P mutant. Scale bars, 5 μm.

(B) The number of CEP152 foci per cell is similar among asynchronous control, V194A, and A199P mutant RPE-1 cell lines. CEP152 localizes to the proximal ends of mother and daughter centrioles, and thus, cells are expected to contain two foci from G1 until centriole disengagement in anaphase. Occasionally, an unfavorable relative orientation, or proximity, prevents resolution of two foci.

(C) Centrin-3 signal is weak in mitotic V194A and A199P mutant RPE-1 cells. Cells arrested in mitosis by monopole-inducing STLC treatment stained by antibodies against the distal centriole marker centrin-3. Images correspond to maximum intensity projections of confocal micrographs. Insets depict high (2×) magnification of centrosomes. Scale bars, 5 μm.

(D) The number of centrin-3 foci per cell is reduced in mitotic V194A mutant RPE-1 cells. Centrin-3 localizes to the distal part of centrioles, and since centriole duplication yields two centriole pairs, cells in mitosis are expected to contain 4 centrioles. Occasionally, the orientation of centrioles within pairs, their proximity, or the small size of nascent centrioles can preclude resolution of pairs, hence the categories 1–2 and 3–4 on the plot.

(E) The number of centrin-3 foci is reduced in asynchronous (non-mitotic) V194A mutant RPE-1 cells with nearly 20% of cells lacking a detectable signal. Asynchronous cells are expected to contain 2–4 centrin-3 foci, depending on their cell-cycle stage.

See also Figures S4–S6.