Mol. Cell. Biol., 01 1998, 536-545, Vol 18, No. 1
Copyright © 1998, American Society for Microbiology

MYC abrogates p53-mediated cell cycle arrest in N-(phosphonacetyl)-L- aspartate-treated cells, permitting CAD gene amplification

OB Chernova, MV Chernov, Y Ishizaka, ML Agarwal and GR Stark
Department of Molecular Biology, Research Institute, The Cleveland Clinic Foundation, Ohio 44195, USA.

Genomic instability, including the ability to undergo gene amplification, is a hallmark of neoplastic cells. Similar to normal cells, "nonpermissive" REF52 cells do not develop resistance to N- (phosphonacetyl)-L-aspartate (PALA), an inhibitor of the synthesis of pyrimidine nucleotides, through amplification of cad, the target gene, but instead undergo protective, long-term, p53-dependent cell cycle arrest. Expression of exogenous MYC prevents this arrest and allows REF52 cells to proceed to mitosis when pyrimidine nucleotides are limiting. This results in DNA breaks, leading to cell death and, rarely, to cad gene amplification and PALA resistance. Pretreatment of REF52 cells with a low concentration of PALA, which slows DNA replication but does not trigger cell cycle arrest, followed by exposure to a high, selective concentration of PALA, promotes the formation of PALA-resistant cells in which the physically linked cad and endogenous N-myc genes are coamplified. The activated expression of endogenous N-myc in these pretreated PALA-resistant cells allows them to bypass the p53-mediated arrest that is characteristic of untreated REF52 cells. Our data demonstrate that two distinct events are required to form PALA-resistant REF52 cells: amplification of cad, whose product overcomes the action of the drug, and increased expression of N-myc, whose product overcomes the PALA-induced cell cycle block. These paired events occur at a detectable frequency only when the genes are physically linked, as cad and N-myc are. In untreated REF52 cells overexpressing N-MYC, the level of p53 is significantly elevated but there is no induction of p21waf1 expression or growth arrest. However, after DNA is damaged, the activated p53 executes rapid apoptosis in these REF52/N-myc cells instead of the long-term protective arrest seen in REF52 cells. The predominantly cytoplasmic localization of stabilized p53 in REF52/N-myc cells suggests that cytoplasmic retention may help to inactivate the growth-suppressing function of p53.

This article has been cited by other articles:

• Karantza-Wadsworth, V., Patel, S., Kravchuk, O., Chen, G., Mathew, R., Jin, S., White, E. (2007). Autophagy mitigates metabolic stress and genome damage in mammary tumorigenesis. Genes Dev. 21: 1621-1635 [Abstract] [Full Text]  
• Ray, S., Atkuri, K. R., Deb-Basu, D., Adler, A. S., Chang, H. Y., Herzenberg, L. A., Felsher, D. W. (2006). MYC Can Induce DNA Breaks In vivo and In vitro Independent of Reactive Oxygen Species.. Cancer Res. 66: 6598-6605 [Abstract] [Full Text]  
• Kandel, E. S., Skeen, J., Majewski, N., Di Cristofano, A., Pandolfi, P. P., Feliciano, C. S., Gartel, A., Hay, N. (2002). Activation of Akt/Protein Kinase B Overcomes a G2/M Cell Cycle Checkpoint Induced by DNA Damage. Mol. Cell. Biol. 22: 7831-7841 [Abstract] [Full Text]  
• El-Hizawi, S., Lagowski, J. P., Kulesz-Martin, M., Albor, A. (2002). Induction of Gene Amplification as a Gain-of-Function Phenotype of Mutant p53 Proteins. Cancer Res. 62: 3264-3270 [Abstract] [Full Text]  
• Felsher, D. W., Zetterberg, A., Zhu, J., Tlsty, T., Bishop, J. M. (2000). Overexpression of MYC causes p53-dependent G2 arrest of normal fibroblasts. Proc. Natl. Acad. Sci. USA 10.1073/pnas.190327097v1 [Abstract] [Full Text]  
• Pise-Masison, C. A., Mahieux, R., Jiang, H., Ashcroft, M., Radonovich, M., Duvall, J., Guillerm, C., Brady, J. N. (2000). Inactivation of p53 by Human T-Cell Lymphotropic Virus Type 1 Tax Requires Activation of the NF-kappa B Pathway and Is Dependent on p53 Phosphorylation. Mol. Cell. Biol. 20: 3377-3386 [Abstract] [Full Text]  
• Ryu, D.-Y., Pratt, V. S. W., Davis, C. D., Schut, H. A. J., Snyderwine, E. G. (1999). In Vivo Mutagenicity and Hepatocarcinogenicity of 2-Amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in Bitransgenic c-myc/{{lambda}}lacZ Mice. Cancer Res. 59: 2587-2592 [Abstract] [Full Text]  
• Felsher, D. W., Bishop, J. M. (1999). Transient excess of MYC activity can elicit genomic instability and tumorigenesis. Proc. Natl. Acad. Sci. USA 96: 3940-3944 [Abstract] [Full Text]  
• Agarwal, M. L., Agarwal, A., Taylor, W. R., Chernova, O., Sharma, Y., Stark, G. R. (1998). A p53-dependent S-phase checkpoint helps to protect cells from DNA damage in response to starvation for pyrimidine nucleotides. Proc. Natl. Acad. Sci. USA 95: 14775-14780 [Abstract] [Full Text]  
• Chernov, M. V., Bean, L. J. H., Lerner, N., Stark, G. R. (2001). Regulation of Ubiquitination and Degradation of p53 in Unstressed Cells through C-terminal Phosphorylation. J. Biol. Chem. 276: 31819-31824 [Abstract] [Full Text]  
• Khan, S. H., Moritsugu, J., Wahl, G. M. (2000). Differential requirement for p19ARF in the p53-dependent arrest induced by DNA damage, microtubule disruption, and ribonucleotide depletion. Proc. Natl. Acad. Sci. USA 97: 3266-3271 [Abstract] [Full Text]  
• Felsher, D. W., Zetterberg, A., Zhu, J., Tlsty, T., Bishop, J. M. (2000). Overexpression of MYC causes p53-dependent G2 arrest of normal fibroblasts. Proc. Natl. Acad. Sci. USA 97: 10544-10548 [Abstract] [Full Text]