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ABSTRACT: The discovery that the single p53 gene encodes several different p53 protein isoforms has initiated a flurry of research into the function and regulation of these novel p53 proteins. Full-length p53 protein level is primarily regulated by the E3-ligase Mdm2, which promotes p53 ubiquitination and degradation. Here, we report that all of the novel p53 isoforms are ubiquitinated and degraded to varying degrees in an Mdm2-dependent and -independent manner, and that high-risk human papillomavirus can degrade some but not all of the novel isoforms, demonstrating that full-length p53 and the p53 isoforms are differentially regulated. In addition, we provide the first evidence that Mdm2 promotes the NEDDylation of p53β. Altogether, our data indicates that Mdm2 can distinguish between the p53 isoforms and modify them differently.
Cell cycle (Georgetown, Tex.) 04/2012; 11(8):1646-55. · 5.36 Impact Factor
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Sergio Menendez, Suzanne Camus,
Aida Herreria,
Ida Paramonov,
Laura B Morera,
Manuel Collado,
Vlad Pekarik,
Iago Maceda,
Michael Edel,
Antonella Consiglio,
Adriana Sanchez,
Han Li,
Manuel Serrano,
Juan C I Belmonte
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ABSTRACT: Embryonic stem (ES) cells and induced pluripotent stem (iPS) cells represent a promising therapeutic tool for many diseases, including aged tissues and organs at high risk of failure. However, the intrinsic self-renewal and pluripotency of ES and iPS cells make them tumorigenic, and hence, the risk of tumor development hinders their clinical application. Here, we present a novel approach to limit their tumorigenicity and increase their safety through increased copy number of tumor suppressors. iPS containing an extra copy of the p53 or Ink4a/ARF locus show normal pluripotency, as determined by in vitro and in vivo differentiation assays. Yet, while retaining full pluripotency, they also possess an improved engagement of the p53 pathway during teratocarcinoma formation, which leads to a reduced tumorigenic potential in various in vitro and in vivo assays. Furthermore, they show an improved response to anticancer drugs, which could aid in their elimination in case tumors arise with no adverse effects on cell function or aging. Our system provides a model for studying tumor suppressor pathways during reprogramming, differentiation, and cell therapy applications. This offers an improved understanding of the pathways involved in tumor growth from engrafted pluripotent stem cells, which could facilitate the use of ES and iPS cells in regenerative medicine.
Aging cell 02/2012; 11(1):41-50. · 7.55 Impact Factor
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ABSTRACT: Embryonic stem (ES) cells are invaluable for their therapeutic potential as well as for the study of early development. Their clinical use demands an understanding of ES cell differentiation, particularly with respect to cell proliferation and the maintenance of genomic integrity, processes for which the transcription factor p53 is essential. However, although the function of p53 as a tumor suppressor has been extensively studied, its role in ES cell biology has not been clearly elucidated. To study p53 activity and regulation in differentiating ES cells, we used knock-in constructs to create a novel reporter system that provides a direct readout of p53 transcriptional activity. We thereby determine that the p53 pathway is active in ES cells, but that p53 activity and the p53-dependent stress response decrease upon differentiation. Although p53 protein levels and activity are usually primarily controlled by the ubiquitin ligase MDM2, we identify the MDM2 homolog MDM4 as the key modulator of p53 activity in differentiating ES cells. Our results provide a better understanding of ES cell regulation and could help to optimize ES cell differentiation protocols for their use in regenerative medicine.
Cell cycle (Georgetown, Tex.) 04/2011; 10(7):1100-8. · 5.36 Impact Factor
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ABSTRACT: The reprogramming of somatic cells to induced pluripotent stem (iPS) cells is one of the major discoveries of recent years. The development and application of patient specific iPS lines could potentially revolutionise cell-based therapy, facilitating the treatment of a wide range of diseases. Despite the numerous technological advancements in the field, an in-depth mechanistical understanding of the pathways involved in reprogramming is still lacking. Several groups have recently provided a mechanistical insight into the role of the p53 tumour suppressor pathway in reprogramming. The repercussions of these findings are profound and reveal an unexpected role of p53 as a "guardian of reprogramming", ensuring genomic integrity during reprogramming at the cost of a reduced efficiency of the process. Here we analyse the latest findings in the field and discuss their relevance for future applications of iPS cell technology.
Cell cycle (Georgetown, Tex.) 10/2010; 9(19):3887-91. · 5.36 Impact Factor
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ABSTRACT: Makorin Ring Finger Protein 1 (MKRN1) is a transcriptional co-regulator and an E3 ligase. Here, we show that MKRN1 simultaneously functions as a differentially negative regulator of p53 and p21. In normal conditions, MKRN1 could destabilize both p53 and p21 through ubiquitination and proteasome-dependent degradation. As a result, depletion of MKRN1 induced growth arrest through activation of p53 and p21. Interestingly, MKRN1 used earlier unknown sites, K291 and K292, for p53 ubiquitination and subsequent degradation. Under severe stress conditions, however, MKRN1 primarily induced the efficient degradation of p21. This regulatory process contributed to the acceleration of DNA damage-induced apoptosis by eliminating p21. MKRN1 depletion diminished adriamycin or ultraviolet-induced cell death, whereas ectopic expression of MKRN1 facilitated apoptosis. Furthermore, MKRN1 stable cell lines that constantly produced low levels of p53 and p21 exhibited stabilization of p53, but not p21, with increased cell death on DNA damage. Our results indicate that MKRN1 exhibits dual functions of keeping cells alive by suppressing p53 under normal conditions and stimulating cell death by repressing p21 under stress conditions.
The EMBO Journal 07/2009; 28(14):2100-13. · 9.20 Impact Factor
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ABSTRACT: The human papillomavirus (HPV) protein E6 can promote the ubiquitination of the p53 tumour suppressor in vitro, providing an explanation for the ability of E6 to induce p53 degradation in vivo and contribute to the potential tumorigenic effect of the virus. Instead, in non-infected cells, p53 levels are primarily destabilised by the ubiquitin E3 ligase activity of the Mdm2 protein. Here we have compared the effects of E6 and Mdm2 on p53 ubiquitination in vivo. We show that whereas in the presence of Mdm2 proteasome inhibitors induce the accumulation of ubiquitinated forms of p53, this does not occur in the presence of E6. Accordingly, we confirm that the effect of E6 and p53 is independent of the six C-terminal lysine residues in p53, which have previously been described to play an important role for effective ubiquitination and degradation of 53 mediated by Mdm2. We also show that other yet unidentified residues in p53 are also susceptible to ubiquitination. These results indicate that E6 does not induce ubiquitination of p53 in the same way as Mdm2 in order to promote its degradation, suggesting important differences between the Mdm2 and E6 effects on p53 degradation.
FEBS Letters 03/2003; 536(1-3):220-4. · 3.54 Impact Factor
