Fig. 5

PACSIN1 functions at the CV stage prior to CP110 loss. a Quantification of CP110 dots in serum starved RPE-1 cells treated with siRNAs for 72 h, serum starved for the last 24 h, and stained with CP110, ^Actub and CEP164 antibodies. CP110 localization on both centrioles (2 dots) or only the daughter centriole (1 dot) was quantified (left panel, siCtrl = 100, siPACS1 = 63 cells). b Quantification of CEP97 dots in cells treated as in a and stained with CEP97, ^Actub, and CEP164 antibodies. CEP97 localization on the centrioles was quantified as in a(siCtrl = 201, siPACS1 = 125 cells). c Quantification of SMOM2-GFP centrosomal accumulation in cells treated as in a, and stained with ^Actub and CEP164 antibodies. “Preciliary” indicates SMOM2-GFP vesicle accumulation at the MC without axonemes (siCtrl = 100, siPACS1 = 67 cells). Scale bar: 2 μm for images in a–c. d Quantification of MC with non-ciliary distal appendages structures from cells treated as in a. Pooled data from 3 independent experiments (siCtrl = 56, siPACS1 = 41 cells). Representative electron micrographs of quantified MCs shown on right. Scale bar: 200 nm. MC mother centriole, DAV distal appendages vesicles, CV ciliary vesicle, or DA non-membrane-associated distal appendages. e–g Quantification of TZ accumulation of TMEM67 (e, siCtrl = 113, siPACS1 = 89 cells), B9D2-GFP (f, siCtrl = 365, siPACS1 = 250 cells), RPGRIP1L (g, siCtrl = 70, siPACS1 = 63 cells) in cells treated as in a. h mIFT20-GFP cells treated as in a, stained with ^Actub and CEP164 antibodies, and quantified for mIFT20 fluorescence intensity at the MC/BB (siCtrl = 46, siPACS1 = 43 cells). Scale bar: 2 μm for images in e–h. In a–c, and e–h, representative images of quantified cells shown on right and means ± SEM, n = 3 independent experiments. Two-tailed t-test analysis as compared with siCtrl was performed. **P < 0.001, ***P < 0.0001