Unexpected functional consequences of xenogeneic transgene expression in beta-cells of NOD mice.

We describe unexpected alterations in the non-obese diabetic (NOD/Lt) mouse model of type 1 diabetes (T1D) following forced beta-cell expression of non-mammalian genes ligated to an insulin promoter sequence. These include the jellyfish green fluorescent protein (GFP), useful for beta-cell identification, and the bacteriophage P1 Cre recombinase, necessary for beta cell-specific ablation of a gene using a Cre-loxP system. Homozygous expression of GFP, driven by the mouse insulin 1 gene promoter (MIP-GFP) in a single transgenic line of NOD mice, produced T1D in postnatal mice that was not associated with insulitis, but rather beta cell-depleted islets. Hemizygous transgene expression suppressed spontaneous autoimmune T1D in females, and produced a male glucose intolerance syndrome associated with age-dependent declines in plasma insulin content. Among lines of transgenic NOD/Lt mice expressing Cre recombinase driven by the rat insulin 2 promoter (RIP-Cre), high, non-mosaic expression correlated with suppressed T1D development. These findings emphasize the need for careful characterization of genetically manipulated NOD mouse stocks to insure that model characteristics have not been compromised.

Tissue and cell-specific expression of the p53-target genes: bax, fas, mdm2 and waf1/p21, before and following ionising irradiation in mice.

The mechanisms by which the p53 tumour suppressor protein would, in vivo, co-ordinate the adaptive response to genotoxic stress is poorly understood. p53 has been shown to transactivate several genes that could be involved in two main cellular responses, growth arrest and apoptosis. To get further insight into the tissue-specific regulation of p53 transcriptional activity, we performed an extensive study looking at the expression of four well characterized p53-responsive genes, before and after gamma-irradiation in p53 wild-type (p53+/+) and p53-deficient (p53-/-) mice. The waf1, bax, fas and mdm2 genes were chosen for their different potential roles in the cellular response to stress. Our data demonstrate the strict p53-dependence of mRNA up-regulation for bax, fas and mdm2 in irradiated tissues and confirm such findings for waf1. They further highlight complex levels of regulatory mechanisms that could lead, in vivo, to selective transcriptional activation of genes by p53. In addition, our results provide arguments for the involvement of p53 in the basal mRNA expression of the four genes in some organs. Finally, in situ expression of Bax and p21Waf-1 protein suggests, at least in lymphoid organs, a direct correlation between selective p53-target gene expression and a particular response of a cell to ionising radiation.

Mechanisms of p53-induced apoptosis: in search of genes which are regulated during p53-mediated cell death.

The tumor suppressor gene p53 is a major player in the protection of cells from DNA damage. In the majority of human cancers, p53 is functionally inactivated--mostly by mutations but also by interaction with viral or cellular proteins. Wild-type p53 is involved in essential functions such as DNA repair, transcription, genomic stability, senescence, cell cycle control and apoptosis. It was shown to be a sequence-specific transcriptional activator, and this activity appears to be necessary to impose growth arrest. A major target gene which participates in p53-mediated growth arrest is p21/Waf1, an inhibitor of cyclin-dependent kinases. Whether or not transcriptional activation of target genes is required for p53-mediated apoptosis may depend on the cell type and external factors, and the mechanism of cell death induction is not clear yet. We have employed clones of the M1 myeloid leukemic cell line expressing a temperature-sensitive p53 mutant to study genes which are regulated during p53-induced apoptosis.