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The p53 protein is a pivotal tumor suppressor that is frequently mutated in many human cancers, although precisely how p53 prevents tumors is still unclear. To add to its complexity, several isoforms of human p53 have now been reported. The Δ133p53 isoform is generated from an alternative transcription initiation site in intron 4 of the p53 gene (Tp53) and lacks the N-terminus. Elevated levels of Δ133p53 have been observed in a variety of tumors. To explore the functions of Δ133p53, we created a mouse expressing an N-terminal deletion mutant of p53 (Δ122p53) that corresponds to Δ133p53. Δ122p53 mice show decreased survival and a different and more aggressive tumor spectrum compared with p53 null mice, implying that Δ122p53 is a dominant oncogene. Consistent with this, Δ122p53 also confers a marked proliferative advantage on cells and reduced apoptosis. In addition to tumor development, Δ122p53 mice show a profound proinflammatory phenotype having increased serum concentrations of interleukin-6 and other proinflammatory cytokines and lymphocyte aggregates in the lung and liver as well as other pathologies. Based on these observations, we propose that human Δ133p53 also functions to promote cell proliferation and inflammation, one or both of which contribute to tumor development.

Murine leukemia viruses (MuLV) induce leukemia through a multistage process, a critical step being the activation of oncogenes through provirus integration. Transcription elements within the long terminal repeats (LTR) are prime determinants of cell lineage specificity; however, the influence of other factors, including the Env protein that modulates cell tropism through receptor recognition, has not been rigorously addressed. The ability of 10A1-MuLV to use both PiT1 and PiT2 receptors has been implicated in its induction of blast cell leukemia. Here we show that restricting receptor usage of 10A1-MuLV to PiT2 results in loss of blast cell transformation capacity. However, the pathogenicity was unaltered when the env gene is exchanged with Moloney MuLV, which uses the Cat1 receptor. Significantly, the leukemic blasts express erythroid markers and consistently contain proviral integrations in the Fli1 locus, a target of Friend MuLV (F-MuLV) during erythroleukemia induction. Furthermore, an NB-tropic variant of 10A1 was unable to induce blast cell leukemia in C57BL/6 mice, which are also resistant to F-MuLV transformation. We propose that 10A1- and F-MuLV actually induce identical (erythro)blastic leukemia by a mechanism involving Fli1 activation and cooperation with inherent genetic mutations in susceptible mouse strains. Furthermore, we demonstrate that deletion of the Icsbp tumor suppressor gene in C57BL/6 mice is sufficient to confer susceptibility to 10A1-MuLV leukemia induction but with altered specificity. In summary, we validate the significance of the env gene in leukemia specificity and underline the importance of a complex interplay of cooperating oncogenes and/or tumor suppressors in determining the pathogenicity of MuLV variants.

Archaea have a eukaryotic type of transcriptional machinery containing homologues of the transcription factors TATA-binding protein (TBP) and TFIIB (TFB) and a pol II type of RNA polymerase, whereas transcriptional regulators identified in archaeal genomes have bacterial counterparts. We describe here a novel regulator of heat shock response, Phr, from the hyperthermophilic archaeon Pyrococcus furiosus that is conserved among Euryarchaeota. The protein specifically inhibited cell-free transcription of its own gene and from promoters of a small heat shock protein, Hsp20, and of an AAA(+) ATPase. Inhibition of transcription was brought about by abrogating RNA polymerase recruitment to the TBP/TFB promoter complex. Phr bound to a 29-bp DNA sequence overlapping the transcription start site. Three sequences conserved in the binding sites of Phr, TTTA at -10, TGGTAA at the transcription start site, and AAAA at position +10, were required for Phr binding and are proposed as consensus regulatory sequences of Pyrococcus heat shock promoters. Shifting the growth temperature from 95 to 103 degrees C caused a dramatic increase of mRNA levels for the aaa(+) atpase and phr genes, but expression of the Phr protein was only weakly stimulated. Our findings suggest that heat shock response in Archaea is negatively regulated by a mechanism involving binding of Phr to conserved sequences.

Detailed knowledge of the immunologic properties of embryonic stem (ES) cells is a prerequisite for safe applications of ES cell-based regenerative medicine. Recently, the long-standing assumption that ES cells are ignored by immunocompetent hosts was disproved. Instead, it is becoming increasingly clear that ES cells actively protect themselves via several immunomodulatory and immunoevasive mechanisms against cytotoxic T-lymphocytes and natural killer cells. Here we review current knowledge about the immunologic properties of ES cells and discuss the implications for ES cell-based regenerative medicine, for the immunobiology of the embryo as a semi-allogeneic graft, and for the regenerative capacity of adult stem cells.

Acute myelogenous leukemia is driven by leukemic stem cells (LSCs) generated by mutations that confer (or maintain) self-renewal potential coupled to an aberrant differentiation program. Using retroviral mutagenesis, we identified genes that generate LSCs in collaboration with genetic disruption of the gene encoding interferon response factor 8 (Irf8), which induces a myeloproliferation in vivo. Among the targeted genes, we identified Mef2c, encoding a MCM1-agamous-deficiens-serum response factor transcription factor, and confirmed that overexpression induced a myelomonocytic leukemia in cooperation with Irf8 deficiency. Strikingly, several of the genes identified in our screen have been reported to be up-regulated in the mixed-lineage leukemia (MLL) subtype. High MEF2C expression levels were confirmed in acute myelogenous leukemia patient samples with MLL gene disruptions, prompting an investigation of the causal interplay. Using a conditional mouse strain, we demonstrated that Mef2c deficiency does not impair the establishment or maintenance of LSCs generated in vitro by MLL/ENL fusion proteins; however, its loss led to compromised homing and invasiveness of the tumor cells. Mef2c-dependent targets included several genes encoding matrix metalloproteinases and chemokine ligands and receptors, providing a mechanistic link to increased homing and motility. Thus, MEF2C up-regulation may be responsible for the aggressive nature of this leukemia subtype.

Mef2c is a MADS (MCM1-agamous–deficient serum response factor) transcription factor best known for its role in muscle and cardiovascular development. A causal role of up-regulated MEF2C expression in myelomonocytic acute myeloid leukemia (AML) has recently been demonstrated. Due to the pronounced monocytic component observed in Mef2c-induced AML, this study was designed to assess the importance of Mef2c in normal myeloid differentiation. Analysis of bone marrow (BM) cells manipulated to constitutively express Mef2c demonstrated increased monopoiesis at the expense of granulopoiesis, whereas BM isolated from Mef2cΔ/− mice showed reduced levels of monocytic differentiation in response to cytokines. Mechanistic studies showed that loss of Mef2c expression correlated with reduced levels of transcripts encoding c-Jun, but not PU.1, C/EBPα, or JunB transcription factors. Inhibiting Jun expression by short-interfering RNA impaired Mef2c-mediated inhibition of granulocyte development. Moreover, retroviral expression of c-Jun in BM cells promoted monocytic differentiation. The ability of Mef2c to modulate cell-fate decisions between monocyte and granulocyte differentiation, coupled with its functional sensitivity to extracellular stimuli, demonstrate an important role in immunity—and, consistent with findings of other myeloid transcription factors, a target of oncogenic lesions in AML.

We report the characterization of TrmB, a protein of 38,800 apparent molecular weight, that is involved in the maltose-specific regulation of a gene cluster in Thermococcus litoralis, malE malF malG orf trmB malK, encoding a binding protein-dependent ABC transporter for trehalose and maltose. TrmB binds maltose and trehalose half-maximally at 20 microm and 0.5 mm sugar concentration, respectively. Binding of maltose but not of trehalose showed indications of sigmoidality and quenched the intrinsic tryptophan fluorescence by 15%, indicating a conformational change on maltose binding. TrmB causes a shift in electrophoretic mobility of DNA fragments harboring the promoter and upstream regulatory motif identified by footprinting. Band shifting by TrmB can be prevented by maltose. In vitro transcription assays with purified components from Pyrococcus furiosus have been established to show pmalE promoter-dependent transcription at 80 degrees C. TrmB specifically inhibits transcription, and this inhibition is counteracted by maltose and trehalose. These data characterize TrmB as a maltose-specific repressor for the trehalose/maltose transport operon of Thermococcus litoralis.