Protein Interaction Between p53 and △113p53 Is Required for the Anti-Apoptotic Function of △113p53

Zebrafish △113p53, an N-terminal truncated p53 isoform, is a p53-target gene that antagonises p53-mediated apoptotic activity.Interestingly, △113p53 does not act on p53 in a dominant-negative manner, but rather interferes with the p53 function by differentially modulating p53-target gene expression to protect cells from apoptosis. Previous studies showed that over-expressed △113p53 and p53proteins formed a complex. However, it is not known whether endogenous p53 and △113p53 proteins also interact with each other, and if this interaction is required for △113p53 to inhibit the apoptotic activity of full-length p53. In this study, we used two available zebrafish p53 antibodies to address these questions. One, Zfp53-N, only recognises full-length p53, whereas the other, Zfp53-A7C10, detects both full-length p53 and △113p53. Using Zfp53-N for immunoprecipitation and Zfp53-A7C10 for detection, we demonstrated that endogenous △113p53 and full-length p53 induced by a DNA-damaging drug formed a complex in vivo. Furthermore, of the six △113p53 mutants we generated with different point mutations in the oligomerisation domain, two failed to interact with p53 and lost the ability to modulate p53-target gene expression and inhibit p53-induced cell apoptosis. However, those △113p53 mutants that could interact with p53 retained the ability to antagonise the apoptotic activity of p53. Therefore, our data demonstrated that proteineprotein interaction between △113p53and p53 is essential for the anti-apoptotic function of △113p53. In addition, the two △113p53 mutants that failed to interact with p53 are also useful for the study of the mechanisms of other functions of △113p53.     

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正Tumour suppressor p53 is activated upon DNA damage stress to promote cell cycle arrest or cell death.An N-terminal truncated p53isoform,zebrafi sh 113p53,is a p53target gene and is strongly induced by DNA damage to inhibit the apoptotic activity of full-length p53.In this issue,Ou et al.report for the fi rst time that endogenous 113p53 and full-length p53 induced by a DNA-damaging drug     

The dichotomy of p53 regulation by noncoding RNAs

The p53 tumor suppressor gene is the most frequently mutated gene in cancer. Significant progress has been made to discern the im- portance of p53 in coordinating cellular responses to DNA damage, oncogene activation, and other stresses. Noncoding RNAs are RNA molecules functioning without being translated into proteins. In this work, we discuss the dichotomy of p53 regulation by noncoding RNAs with four unconventional questions. First, is overexpression of microRNAs responsible for p53 inactivation in the absence of p53 mutation？ Second, are there somatic mutations in the noncoding regions of the p53 gene？ Third, is there a germline mutant in the non- coding regions of the p53 gene that predisposes carriers to cancer？ Fourth, can p53 activation mediated by a noncoding RNA mutation cause cancer？ This work highUghts the prominence of noncoding RNAs in p53 dysregutation and tumorigenesis.     

Differential effects of p63 mutants on transactivation of p53 and/or p63 responsive genes

p63, known to play a role in development, has more recently also been implicated in cancer progression. Mutations in p63 have been shown to be responsible for several human developmental diseases. Differential splicing of the p63 gene gives rise to p63 isoforms, which can act either as tumor suppressors or as oncogene. In this report, we studied the effects of naturally occurring TAp637 mutants on the regulation of p53/p63 and p63 specific target genes. We observed significant differences among p63 mutants to regulate the p53/p63 and p63 specific target genes. Additionally, we observed a differential effect of p63 mutants on wildtype-p63-mediated induction ofp53/p63 and p63 specific target genes. We also demonstrated that these mutants differentially regulate the binding of wildtype p63 to the promoter of target genes. Furthermore, the effects of these mutants on cell death and survival were consistent with their ability to regulate the downstream targets when compared to wildtype TAp63T. In summary, our data demonstrate that p63 mutants exhibit differential effects on p63 and p53/p63 specific target genes and on the induction of apoptosis, and provide further insight into the function of p63.     

Knockdown of human p53 gene expression in 293-T cells by retroviral vector-mediated short hairpin RNA

RNA interference （RNAi） is an evolutionarily conserved process of gene silencing in multiple organisms, which has become a powerful tool for investigating gene function by reverse genetics. Recently, many groups have reported to use synthesized oligonucleotides or siRNA encoding plasmids to induce RNAi in mammalian cells by transfection, but this is still limited in its application, especially when it is necessary to generate long-term gene silencing in vivo. To circumvent this problem, retrovirus- or lentivirus-delivered RNAi has been developed. Here, we described two retroviral systems for delivering short hairpin RNA （shRNA） transcribed from the H1 promoter. The results showed that retroviral vector-mediated RNAi can substantially downregulate the expression of human p53 in 293-T cells. Furthermore, the retroviral vectormediated RNAi in our transduction system can stably inactivate the p53 gene for a long time. Compared to shRNAs transcribed from the U6 promoter, HI-driven shRNA also dramatically reduced the expression of p53. The p53 downregulation efficiencies of H1- and U6-driven shRNAs were almost identical. The results indicate that retroviral vector-delivered RNAi would be a useful tool in functional genomics and gene therapy.     

Genetic Analysis and Mapping of the Dominant Dwarfing Gene D-53 in Rice

The dwarfing gene D-53 Is one of a few dominant genes for dwarfing In rice （Oryza satlva L.）. In the present study, our genetic analysis confirmed that mutant characteristics including dwarfing, profuse tlllerlng, thin stems and small panicles are all controlled by the dominant D-53 gene. We measured the length of each Internode of KL908, a D-53-carrylng line, and classified the dwarfism of KL908 Into the dn-type. In addition, we measured elongation of the second sheath and a-amylase activity In the endosperm, and we characterized KL908 as a dwarf mutant that was neither glbberelllc acid-deficient nor glbberelllc acid-Insensitive. Using a large F2 population obtained by crossing KL908 with a wild-type variety, NJ6, the D-53 gene was mapped to the terminal region of the short arm of chromosome 11, with one simple sequence repeat marker, Ds3, co-segregating, and the other, K81114, located 0.6 cM away.     

Oncoprotein p28^GANK binds to RelA and retains NF-κB in the cytoplasm through nuclear export

p28^GANK （also known as PSMD 10, p28 and gankyrin） is an ankyrin repeat anti-apoptotic oncoprotein that is commonly overexpressed in hepatocellular carcinomas and increases the degradation of p53 and Rb. NF-IκB （nuclear factor-κB） is known to be sequestered in the cytoplasm by IκB （inhibitor of NF-κB） proteins [1, 2], but much less is known about the cytoplasmic retention of NF-κB by other cellular proteins. Here we show that p28^GANK inhibits NF-κB activity. As a nuclear-cytoplasmic shuttling protein, p28^GANK directly binds to NF-κB/RelA and exports RelA from nucleus through a chromosomal region maintenance-1 （CRM-1） dependent pathway, which results in the cytoplasmic retention of NF- κB/RelA. We demonstrate that all the ankyrin repeats of p28^GANK are required for the interaction with RelA and that the N terminus of p28^GANK, which contains the nuclear export sequence （NES）, is responsible for suppressing NF-κB/RelA nuclear translocation. These results suggest that overexpression of p28^GANK prevents the nuclear localization and inhibits the activity of NF-κB/RelA.     

Moderate hypothermia prevents neural cell apoptosis following spinal cord ischemia in rabbits

Paraplegia is a disastrous complication after operations of descending and thoracoabdominal aortic aneurysm. Regional hypothermia protects against spinal cord ischemia although the protective mechanism is not well know. The objective of this study is to examine whether hypothermia protects the spinal cord by preventing apoptosis of nerve cell and also investigate a possible mechanism involved in hypothermia neuroprotection. Cell apoptosis with necrosis was evident in the spinal cord 24 h after 30 min of ischemia. Moderate hypothermia decreased the incidence of apoptotic nerve cells. Both cell apoptosis and necrosis were attenuated by hypothermia, p53 expression increased and bcl-2 expression declined after ischemia, while hypothermia mitigated these changes. This study suggests that apoptosis contributes to cell death after spinal cord ischemia, and that moderate hypothermia can prevent nerve cell apoptosis by a mechanism associated with bcl-2 and p53 genes.     

Role ofthe IncRNA-p53 regulatory network in cancer

Advances in functional genomics have led to discovery of a large group of previous uncharacterized long non-coding RNAs （IncRNAs）. Emerging evidence indicates that IncRNAs may serve as master gene regulators through various mechanisms. Dysregulation of IncRNAs is often associated with a variety of human diseases including cancer. Of significant interest, recent studies suggest that IncRNAs participate in the p53 tumor suppressor regulatory network. In this review, we discuss how IncRNAs serve as p53 regulators or p53 effectors. Further characterization of these p53-associated IncRNAs in cancer will provide a better understanding of lncRNA- mediated gene regulation in the p53 pathway. As a result, IncRNAs may prove to be valuable biomarkers for cancer diagnosis or poten- tial targets for cancer therapy.     

Genetic aberration in primary hepatocellular carcinoma：correlation between p53 gene mutation and loss—of—heterozygosity on chromosome 16q21—q23 and 9p21—p23

To elucidate the molecular pathology underlying the development of hepatocellular carcinoma (HCC),we used 41 highly polymorphic microsatellite markers to examine 55 HCC and corresponding non-tumor liver tissues on chromosome 9,16 and 17.Loss-of-heterozygosity(LOH) is observed with high frequency on chromosomal region 17p13(36k/55,65%),9q21-p23(28/55,51%),16q21-23(27/55,49%) in tumors.Meanwhile,microsatellite instability is rarely found in these microsatellite loci.Direct sequencing was performed to detect the tentative mutation of tumor wuppressor genes in these regions:p53,MTS1/p16,and CDH1/E-cadherin.Wihin exon 5-9 of p53 gene,14 out of 55 HCC specimens(24%) have somatic mutations,and nucleotide deletion of this gene is reported in HCC for the first time.Mutation in MTS1/p16 is found only in one tumor case.We do not find mutations in CDH1/E-cadherin.Furthermore,a statistically significant correlation is present between p53 gene mutation and loss of chromosome region 16q21-q23 and 9p21-p23,which indicates that synergism between p53 inactivation and deletion of 16q21-q23 and 9p21-p23 may play a role in the pathogenesis of HCC.     

Induction of antiproliferative effect by diosgenin through activation of p53,release of apoptosis-inducing factor （AIF） and modulation of caspase-3 activity in different human cancer cells

Previously, we demonstrated that a plant steroid, diosgenin, altered cell cycle distribution and induced apoptosis in the human osteosarcoma 1547 cell line. The objective of this study was to investigate if the antiproliferative effect of diosgenin was similar for different human cancer cell lines such as laryngocarcinoma HEp-2 and melanoma M4Beu cells. Moreover, this work essentially focused on the mitochondrial pathway. We found that diosgenin had an important and similar antiproliferative effect on different types of cancer cells. In addition, our new results show that diosgenin-induced apoptosis is caspase-3 dependent with a fall of mitochondrial membrane potential, nuclear localization of AIF and poly (ADP-ribose) polymerase cleavage. Diosgenin treatment also induces p53 activation and cell cycle arrest in the different cell lines studied.