Regulation of Apoptosis and Differentiation by p53 in Human ...

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CHAPTER 1: Introduction (Jones et al., 1995; Montes de Oca Luna et al., 1995). This implies that at an early embryonic stage p53 is functional and its functional activity has to be controlled to allow survival of embryos and subsequent development. Embryonic phenotypes of homozygotes deficient for either gene appear more obvious upon challenge. Lack of either of these genes is sufficient to rescue blastocysts from cell death induced by hyperglycaemia (Keim et al., 2001; Moley et al., 1998). Suppression of hyperglycaemia-induced cell death is dependent on both Bax and p53, and is accompanied by a decrease in the number of embryo resorptions, directly implicating cell death in postimplantation embryonic loss (Moley, 2001). 1.4.2- p53 in apoptosis There are different apoptotic pathways controlled by p53. In response to cellular stress, p53 transactivates proapoptotic genes (PUMA, Noxa, Bax, p53 AIP1, Apaf-1 and PERP) activating the receptor-mediator (extrinsic) pathway, the mitochondrial (intrinsic) pathway and the ER pathway (Figure 1.4). p53 can also repress the transcription of relevant prosurvival genes (Bcl2 and IAPs). Bioinformatic studies predict that more than 4000 human genes contain putative p53 binding sites (Lu, 2005). The activity of p53 is controlled in different ways depending on the cell context either trough post-translational modifications (phosphorylation, sumoylation, acetylation) or through the activity of cofactors and inhibitors, like Mdm2 (Bourdon et al., 2003). Several in vivo studies reported that a fraction of induced p53 translocates to the mitochondria. A study published by Erster et al (Erster et al., 2004) showed that in sensitive organs, like the thymus, mitochondrial p53 accumulation in vivo occurs soon after a death stimulus, triggering a rapid first wave of apoptosis that is transcription independent and may precede a second slower wave that is transcription dependent. At the mitochondrion p53 interacts with Bcl2 family of regulators of mitochondrial permeability (Chipuk et al., 2004; Leu et al., 2004). - 28 -
CHAPTER 1: Introduction Mitochondrion Death receptor Bad Bax Bid Adaptor Cyt-c Bcl2 Bid Casp-8 10 PUMA Cyt-c Casp-9 Apaf-1 DNA damage Apoptosome Casp-3 p53 Post-translational modifications Bax Puma Noxa GADD45 p21 MDM2 Cleavage of substrates Nucleus APOPTOSIS p14ARF FIGURE 1.4- Schematic representation of apoptotic pathways controlled by p53. DNA damage and other stress signals converge on p53 signaling by activation of protein kinases and/or acetyltransferases, which phosphorylate or acetylate p53, respectively. The inactivation of p53 by Mdm2 is counteracted by mechanisms involving post-translational modification of p53 or the Mdm2 inhibitor ARF. These posttranslational modifications generally result not only in stabilization, but also in activation of p53 in the nucleus, where p53 interacts with sequencespecific DNA binding sites in its target gene promoters. Through its activity as a sequencespecific transcriptional activator, p53 increases the expression of pro-apoptotic Bcl-2 family proteins. p53 can also have a direct apoptogenic role on the mitochondria, interacting with Bcl2 family of regulators of mitochondrial permeability. Released apoptogenic factors facilitate the activation of the effector caspases through the Apaf-1–caspase 9-apoptosome. p53 can be activated pharmacologically by the topoisomerase II inhibitor etoposide. This is an antineoplastic drug that has been widely used to induced apoptosis in response to DNA damage (Karpinich et al., 2002). Topoisomerase II is a nuclear enzyme that functions during both DNA replication and transcription (Hande, 1998). Topoisomerase II prevents "knots" from forming in DNA by allowing the passage of an intact segment of the helical DNA through a transient double strand break (Burden and Osheroff, 1998). Topoisomerase II inhibitors such as etoposide stabilize the complex formed by topoisomerase II and the 5'-cleaved ends of the DNA, thus forming stable (nonrepairable) protein-linked DNA double strand breaks (Burden and Osheroff, 1998). Cells are apparently able to recognize such DNA damage and, in turn, to eliminate the injured cells by apoptosis. This is known to be p53-mediated via the mitochondrial death pathway. - 29 -
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CHAPTER 3: Characterization of hESC
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CHAPTER 6: Bibliography Benard, J.,
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CHAPTER 6: Bibliography Duensing, A
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CHAPTER 6: Bibliography Melino, G.,
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CHAPTER 6: Bibliography Wei, C. L.,
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APPENDIX I - 144 -
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APPENDIX I 4% Paraformaldehyde (PFA
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TABLE A 2.1- List of genes upregula
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PMP22 1.76 0.70 NM_153322 PDE6G 1.8
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TABLE A 2.2- List of functions of u
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Protein Synthesis Ophthalmic Diseas
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1.73 0.897 CN292254 LOC651029 1.73
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Reproductive System Disease Cellula
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