Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2015 Jun 22;209(6):863–878. doi: 10.1083/jcb.201502088

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© 2015 Moyer et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

PMC Copyright notice

Figure 1. — Plk4 kinase activity is required to maintain STIL at the centriole. (A) Schematic of the strategy used to knock-in the AS mutation into both alleles of Plk4 in human DLD-1 cells. The repair oligonucleotide introduced the AS mutation (L89G), a silent AflIII restriction site, and a mutation in the protospacer adjacent motif (PAM) to prevent recutting by SpCas9 after homology-directed repair. (B) Plk4^WT/WT or Plk4^AS/AS cells were treated with 3MB-PP1 for 20 h and nocodazole was added for the final 4 h of the treatment. The graph shows the fraction of mitotic cells with the indicated number of centrioles. Bars represent the mean of three independent experiments, with >20 cells counted per experiment. (C) Selected images of mitotic Plk4^AS/AS cells from B stained with Centrin and CEP192. Bars: (large images) 5 µm; (inset images) 0.5 µm. (D) Quantification of the relative levels of Plk4 at the centrosome of interphase cells 20 h after addition of 3MB-PP1. Bars represent the mean of at least three independent experiments, with >40 cells counted per experiment. (E) Graph showing the increase in cell number at various times after addition of 3MB-PP1. Points show the mean of at least three independent experiments. (F) Quantification of relative protein abundance at the centrosome of S/G2 phase cells 1 h after the addition of 3MB-PP1. Bars represent the mean of three independent experiments, with >40 cells counted per experiment. (G) Immunoblot showing no change in the level of endogenous STIL and SAS6 at 1 or 2 d after Plk4 inhibition with 3MB-PP1. All error bars represent the SEM.