Figure 2. Phosphomimetic STIL Supports Centriole Assembly but Cannot Bypass the Requirement of PLK4.

(A–C) Acentriolar PLK4^as; STIL^−/− cells stably carrying exogenous, tetracycline-inducible STIL constructs were induced to express STIL^WT (A), STIL^4A (B), or STIL^4D (C) in the presence or absence of 3MB-PP1, as indicated, and examined for centriole assembly using SAS-6, STIL, and centrin antibodies. Note that STIL^WT (A) and STIL^4D (C) could drive centriole assembly only when PLK4 was active. STIL^4A is not functional in either case (B). Error bars represents SD; n > 25, N = 3.

(D and E) The efficiency of centriole duplication during S phase (BrdU) was examined in stable PLK4^asKI; STIL^{WT or 4D} cell lines in which centriole biogenesis is supported by exogenously expressed STIL^WT or STIL^4D, using indicated antibodies (D). Note that centrioles duplicate normally in most STIL^WT and STIL^4D cells under low DOX, forming centriole doublets each labeled with 1 STIL dot (1 STIL/Doublet). A minor centriole over-duplication is noted in ~7.5% of STIL^4D cells (E, green), forming centriole triplets or rosettes each labeled with more than one STIL dots (> 1 STIL/overduplication). Error bars represent SD; n > 50, N = 3.