Overexpression of mutant p53 is a common theme in tumors, suggesting a selective pressure for p53 mutation in cancer development and progression. To determine how mutant p53 expression may lead to survival advantage in human cancer cells, we generated stable cell lines expressing p53 mutants p53-R175H, -R273H, and -D281G by use of p53-null human H1299 (lung carcinoma) cells. Compared to vector-transfected cells, H1299 cells expressing mutant p53 showed a survival advantage when treated with etoposide, a common chemotherapeutic agent; however, cells expressing the transactivation-deficient triple mutant p53-D281G (L22Q/W23S) had significantly lower resistance to etoposide. Gene expression profiling of cells expressing transcriptionally active mutant p53 proteins revealed the striking pattern that all three p53 mutants induced expression of approximately 100 genes involved in cell growth, survival, and adhesion. The gene NF-B2 is a prominent member of this group, whose overexpression in H1299 cells also leads to chemoresistance. Treatment of H1299 cells expressing p53-R175H with small interfering RNA specific for NF-B2 made these cells more sensitive to etoposide. We have also observed activation of the NF-B2 pathway in mutant p53-expressing cells. Thus, one possible pathway through which mutants of p53 may induce loss of drug sensitivity is via the NF-B2 pathway.Mutation in the p53 tumor suppressor gene is a common event in human cancer (6,36,40,43,44,65,71,72). Unlike what is seen for other tumor suppressors, in the majority of human carcinomas with p53 mutations, a protein with one amino acid substitution is overexpressed, suggesting the existence of a selection pressure for maintaining expression of the mutant protein (6,36,40,43,44,65,71,72). This also is perhaps indicative of an active role played by p53 mutants in oncogenesis and follows the gain-of-function hypothesis, which predicts not only that mutations in the p53 gene destroy the tumor suppressor function of the wild-type (WT) protein but that the mutant proteins may also gain oncogenic functions. The gain-of-function hypothesis also predicts that tumors with mutant p53 proteins may be more aggressive or that patients with tumors harboring mutant p53 have poorer prognoses than patients with tumors lacking the p53 protein. This has been found to be true for various types of cancers (10,28,29,81,82,88).WT p53 is a sequence-specific transactivator of promoters containing p53-binding sites. Elevated levels of WT p53 in response to cellular stress situations, such as DNA damage, can lead to apoptosis or induce cell cycle arrest (26,53,60,68,84,89,91) by inducing expression of genes involved in various aspects of cellular growth regulation (21,26,27,49,50,53,60,63,68,84,89,91). A mutation in one allele of p53 generates a stable mutant protein with compromised tumor suppressor function. However, there is compelling evidence to suggest that apart from loss of growth suppressor function, p53 mutants can confer oncogenic properties even in the absence of WT p53...

ABSTRACTp53 mutants with a single amino acid substitution are overexpressed in a majority of human cancers containing a p53 mutation. Overexpression of the mutant protein suggests that there is a selection pressure on the cell indicative of an active functional role for mutant p53. Indeed, H1299 cells expressing mutant p53-R175H, p53-R273H or p53-D281G grow at a faster rate compared with a control cell line. Using p53-specific small interfering RNA, we show that the growth rate of mutant p53-expressing cells decreases as mutant p53 level decreases, demonstrating that the increased cellular growth is dependent on p53 expression. Increased growth rate is not observed for H1299 cell clones expressing mutant p53-D281G (L22Q/ W23S), which has been shown to be defective in transactivation in transient transcriptional assays. This shows that the increased growth rate imparted by mutant p53 in H1299 cells requires the transactivation function of mutant p53. By performing microarray hybridization analyses, we show that constitutive expression of three common p53 mutants (p53-R175H, p53-R273H, and p53-D281G) in H1299 human lung carcinoma cells evokes regulation of a common set of genes, a significant number of which are involved in cell growth regulation. Predictably, H1299 cells expressing p53-D281G (L22Q/W23S) are defective in upregulating a number of these genes. The differences in expression profiles induced by individual p53 mutants in the cells may be representative of the p53 mutants and how they can affect gene expression resulting in the observed "gain of function" phenotypes (i.e., increased growth rate, decreased sensitivity to chemotherapeutic agents, and so forth).

Alkaloid; cytochrome P450; gene silencing; nicotine N-demethylase; N'-nitrosonornicotine; plant genetic engineering; metabolic engineering; Nicotiana tabacum L.; real-time PCR; RNA interference; tobacco-specific nitrosamines.

Background: IRF7 is known as the master regulator of type I IFN production, yet little is known about its negative regulation. Results: AIP interacts with IRF7 and inhibits IRF7 nuclear localization. Conclusion: AIP suppresses virus-induced type I IFN production by targeting IRF7 for inactivation. Significance: Understanding IRF7 regulation is important to elucidate the host innate immune response to virus infection.