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. 2009 Mar 23;29(10):2582–2593. doi: 10.1128/MCB.00952-08

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Copyright © 2009, American Society for Microbiology

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FIG. 5. — Loss of p53 improves growth of Cdk2⁻^/⁻ Cdk4⁻^/⁻ cells. (A) 3T3 assay performed for 16 passages using WT, Cdk2⁻^/⁻ Cdk4⁻^/⁻, Cdk2⁻^/⁻ Cdk4⁻^/⁻ p53⁻^/⁻, Cdk2⁻^/⁻ Cdk4⁻^/⁻ p53^+/⁻, and p53⁻^/⁻ MEFs. The x axis shows the number of passages, and the y axis indicates the cumulative cell number for each passage. (B) Western blot (WB) analysis. One hundred micrograms of protein lysate from passage 3 MEFs of WT (lane 1), Cdk2⁻^/⁻ Cdk4⁻^/⁻ (lane 2), Cdk2⁻^/⁻ Cdk4⁻^/⁻ p53⁻^/⁻ (lane 3), and p53⁻^/⁻ (lane 4) genotypes was separated on a 12.5% gel, and proteins were transferred to a polyvinylidene difluoride membrane. Antibodies against Hsp90, Cdk2, Cdk4, p53, p21, and p27 were used for detection. (C to F) β-Galactosidase staining. Shown are representative images from WT (C), Cdk2⁻^/⁻ Cdk4⁻^/⁻ (D), p53⁻^/⁻ (E), and Cdk2⁻^/⁻ Cdk4⁻^/⁻ p53⁻^/⁻ (F) MEFs stained with β-galactosidase as a marker for senescence. (G) Graph depicting the percentage of proliferating cells and senescent cells at passage 3 in WT, Cdk2⁻^/⁻ Cdk4⁻^/⁻, Cdk2⁻^/⁻ Cdk4⁻^/⁻ p53⁻^/⁻, and p53⁻^/⁻ MEFs, counting at least 200 cells for each genotype.

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