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p53 protects against genome instability following centriole duplication failure.

Lambrus BG, Uetake Y, Clutario KM, Daggubati V, Snyder M, Sluder G, Holland AJ - J. Cell Biol. (2015)

Bottom Line: Depleting p53 allowed cells that fail centriole duplication to proliferate indefinitely.Washout of auxin and restoration of endogenous Plk4 levels in cells that lack centrioles led to the penetrant formation of de novo centrioles that gained the ability to organize microtubules and duplicate.In summary, we uncover a p53-dependent surveillance mechanism that protects against genome instability by preventing cell growth after centriole duplication failure.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205.

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The centriole localization of STIL requires Plk4. (A) Quantification of Plk4 protein levels at the centrosome of interphase cells at 24 h after addition of the indicated concentrations of IAA (gray bars). The fraction of mitotic cells that underwent successful centriole duplication (three or more centrioles) was quantified in the same samples (open bars). Bars represent the mean of >40 cells from two independent experiments. (B) Representative images of Plk4AID/AID cells immunostained with CEP192 and Centrin after treatment with the indicated dose of IAA for 24 h. Bars: (main) 5 µm; (inset) 1 µm. (C) Quantification of relative protein abundance at the centrosome of S or G2 phase (CENP-F positive) cells 1 h after IAA addition. Centriole recruitment of STIL requires Plk4. r-SAS6, rabbit SAS6 antibody; m-SAS6, mouse SAS6 antibody. Bars represent the mean of >35 cells from two independent experiments. Immunoblot shows no change in the level of endogenous STIL after Plk4 degradation. All error bars in the figure represent the SEM.
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fig2: The centriole localization of STIL requires Plk4. (A) Quantification of Plk4 protein levels at the centrosome of interphase cells at 24 h after addition of the indicated concentrations of IAA (gray bars). The fraction of mitotic cells that underwent successful centriole duplication (three or more centrioles) was quantified in the same samples (open bars). Bars represent the mean of >40 cells from two independent experiments. (B) Representative images of Plk4AID/AID cells immunostained with CEP192 and Centrin after treatment with the indicated dose of IAA for 24 h. Bars: (main) 5 µm; (inset) 1 µm. (C) Quantification of relative protein abundance at the centrosome of S or G2 phase (CENP-F positive) cells 1 h after IAA addition. Centriole recruitment of STIL requires Plk4. r-SAS6, rabbit SAS6 antibody; m-SAS6, mouse SAS6 antibody. Bars represent the mean of >35 cells from two independent experiments. Immunoblot shows no change in the level of endogenous STIL after Plk4 degradation. All error bars in the figure represent the SEM.

Mentions: To examine whether treatment with IAA leads to the expected failure of centriole duplication, we assessed centriole number in cells undergoing mitosis one cell cycle (30 h) after IAA addition. In untreated Plk4AID/AID cells, centriole duplication occurred successfully in >90% of cells (0 µM IAA; Fig. 2, A and B). In contrast, IAA addition caused a failure of centriole duplication in >90% of Plk4AID/AID cells (500 µM IAA; Fig. 2, A and B). We conclude that Plk4AID/AID cells offer a new tool to achieve rapid and reversible depletion of endogenous Plk4.


p53 protects against genome instability following centriole duplication failure.

Lambrus BG, Uetake Y, Clutario KM, Daggubati V, Snyder M, Sluder G, Holland AJ - J. Cell Biol. (2015)

The centriole localization of STIL requires Plk4. (A) Quantification of Plk4 protein levels at the centrosome of interphase cells at 24 h after addition of the indicated concentrations of IAA (gray bars). The fraction of mitotic cells that underwent successful centriole duplication (three or more centrioles) was quantified in the same samples (open bars). Bars represent the mean of >40 cells from two independent experiments. (B) Representative images of Plk4AID/AID cells immunostained with CEP192 and Centrin after treatment with the indicated dose of IAA for 24 h. Bars: (main) 5 µm; (inset) 1 µm. (C) Quantification of relative protein abundance at the centrosome of S or G2 phase (CENP-F positive) cells 1 h after IAA addition. Centriole recruitment of STIL requires Plk4. r-SAS6, rabbit SAS6 antibody; m-SAS6, mouse SAS6 antibody. Bars represent the mean of >35 cells from two independent experiments. Immunoblot shows no change in the level of endogenous STIL after Plk4 degradation. All error bars in the figure represent the SEM.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4494000&req=5

fig2: The centriole localization of STIL requires Plk4. (A) Quantification of Plk4 protein levels at the centrosome of interphase cells at 24 h after addition of the indicated concentrations of IAA (gray bars). The fraction of mitotic cells that underwent successful centriole duplication (three or more centrioles) was quantified in the same samples (open bars). Bars represent the mean of >40 cells from two independent experiments. (B) Representative images of Plk4AID/AID cells immunostained with CEP192 and Centrin after treatment with the indicated dose of IAA for 24 h. Bars: (main) 5 µm; (inset) 1 µm. (C) Quantification of relative protein abundance at the centrosome of S or G2 phase (CENP-F positive) cells 1 h after IAA addition. Centriole recruitment of STIL requires Plk4. r-SAS6, rabbit SAS6 antibody; m-SAS6, mouse SAS6 antibody. Bars represent the mean of >35 cells from two independent experiments. Immunoblot shows no change in the level of endogenous STIL after Plk4 degradation. All error bars in the figure represent the SEM.
Mentions: To examine whether treatment with IAA leads to the expected failure of centriole duplication, we assessed centriole number in cells undergoing mitosis one cell cycle (30 h) after IAA addition. In untreated Plk4AID/AID cells, centriole duplication occurred successfully in >90% of cells (0 µM IAA; Fig. 2, A and B). In contrast, IAA addition caused a failure of centriole duplication in >90% of Plk4AID/AID cells (500 µM IAA; Fig. 2, A and B). We conclude that Plk4AID/AID cells offer a new tool to achieve rapid and reversible depletion of endogenous Plk4.

Bottom Line: Depleting p53 allowed cells that fail centriole duplication to proliferate indefinitely.Washout of auxin and restoration of endogenous Plk4 levels in cells that lack centrioles led to the penetrant formation of de novo centrioles that gained the ability to organize microtubules and duplicate.In summary, we uncover a p53-dependent surveillance mechanism that protects against genome instability by preventing cell growth after centriole duplication failure.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205.

Show MeSH
Related in: MedlinePlus