The prion hypothesis posits that a misfolded form of prion protein (PrP) is responsible for the infectivity of prion disease. Using recombinant murine PrP purified from Escherichia coli, we created a recombinant prion with the hallmarks of the pathogenic PrP isoform: aggregated, protease-resistant, and self-perpetuating. After intracerebral injection of the recombinant prion, wild-type mice developed neurological signs in ~130 days and reached the terminal stage of disease in ~150 days. Characterization of diseased mice revealed classic neuropathology of prion disease, the presence of protease-resistant PrP, and the capability of serially transmitting the disease, confirming that these mice succumbed to prion disease. Thus, as postulated by the prion hypothesis, the infectivity in mammalian prion disease results from an altered conformation of PrP.

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The Forkhead Box (Fox) proteins are an extensive family of transcription factors that shares homology in the winged helix DNAbinding domain and whose members play essential roles in cellular proliferation, differentiation, transformation, longevity, and metabolic homeostasis. Liver regeneration studies with transgenic mice demonstrated that FoxM1B regulates the onset of hepatocyte DNA replication and mitosis by stimulating expression of cell cycle genes. Here, we demonstrate that albumin-promoter-driven Cre recombinase-mediated hepatocyte-specific deletion of the Foxm1b Floxed (fl) targeted allele resulted in significant reduction in hepatocyte DNA replication and inhibition of mitosis after partial hepatectomy. Reduced DNA replication in regenerating Foxm1b ؊/؊ hepatocytes was associated with sustained increase in nuclear staining of the cyclin-dependent kinase (Cdk) inhibitor p21 Cip1 (p21) protein between 24 and 40 h after partial hepatectomy. Furthermore, increased nuclear p21 levels and reduced expression of Cdc25A phosphatase coincided with decreases in Cdk2 activation and hepatocyte progression into S-phase. Moreover, the significant reduction in hepatocyte mitosis was associated with diminished mRNA levels and nuclear expression of Cdc25B phosphatase and delayed accumulation of cyclin B1 protein, which is required for Cdk1 activation and entry into mitosis. Cotransfection studies demonstrate that FoxM1B protein directly activated transcription of the Cdc25B promoter region. Our present study shows that the mammalian Foxm1b transcription factor regulates expression of cell cycle proteins essential for hepatocyte entry into DNA replication and mitosis.knock-out mouse ͉ Cdc25A ͉ Cdc25B ͉ cyclin-dependent kinase inhibitor p21 Cip1

Prions containing misfolded prion protein (PrP Sc ) can be formed with cofactor molecules using the technique of serial protein misfolding cyclic amplification. However, it remains unknown whether cofactors materially participate in maintaining prion conformation and infectious properties. Here we show that withdrawal of cofactor molecules during serial propagation of purified recombinant prions caused adaptation of PrP Sc structure accompanied by a reduction in specific infectivity of >10 5 -fold, to undetectable levels, despite the ability of adapted "protein-only" PrP Sc molecules to self-propagate in vitro. We also report that changing only the cofactor component of a minimal reaction substrate mixture during serial propagation induced major changes in the strain properties of an infectious recombinant prion. Moreover, propagation with only one functional cofactor (phosphatidylethanolamine) induced the conversion of three distinct strains into a single strain with unique infectious properties and PrP Sc structure. Taken together, these results indicate that cofactor molecules can regulate the defining features of mammalian prions: PrP Sc conformation, infectivity, and strain properties. These findings suggest that cofactor molecules likely are integral components of infectious prions.phospholipid | bioassay | repertoire | convergence | diversity

Conditional deletion of the mouse Forkhead Box (Fox) m1b targeted allele in adult hepatocytes (Foxm1, previously called HFH-11B, Trident, Win, or MPP2) demonstrated that the Foxm1b transcription factor is essential for hepatocyte mitosis during liver regeneration. To determine the role of Foxm1b in liver development, we have generated Foxm1b -/- mice that deleted the Foxm1b exons encoding the winged helix DNA binding and transcriptional activation domains. Here, we show that all of the Foxm1b -/- embryos died in utero by 18.5 days postcoitum (dpc). Embryonic Foxm1b -/- livers displayed a 75% reduction in the number of hepatoblasts, resulting from diminished DNA replication and a failure to enter mitosis causing a polyploid phenotype. Reduced hepatoblast mitosis was associated with decreased protein levels of the Polo-like kinase 1 and Aurora B kinase, which phosphorylate regulatory proteins essential for orchestrating mitosis and cytokinesis. Diminished proliferation of Foxm1b -/- hepatoblasts contributed to abnormal liver development with significant reduction in the number of large hepatic veins compared to embryonic wild-type (WT) liver. Furthermore, embryonic Foxm1b -/- livers did not develop intrahepatic bile ducts, and these presumptive biliary hepatoblasts failed to express either biliary cytokeratins or nuclear levels of hepatocyte nuclear factor 1beta. These results suggest that Foxm1b is critical for hepatoblast precursor cells to differentiate toward biliary epithelial cell lineage. Finally, we used a hepatoblast-specific Cre recombinase transgene to mediate deletion of the Foxm1b fl/fl allele in the developing liver, and these embryos died in utero and exhibited diminished hepatoblast proliferation with similar abnormalities in liver morphogenesis, suggesting that the defect in liver development contributed to embryonic lethality.

The forkhead box (Fox) family of transcription factors share homology in the winged helix͞forkhead DNA-binding domain and play important roles in regulating cellular proliferation, differentiation, longevity, and cellular transformation. Forkhead box M1B (FoxM1B) is a ubiquitously expressed member of the Fox transcription factor family whose expression is restricted to proliferating cells and that mediates hepatocyte entry into DNA synthesis and mitosis during liver regeneration. Recent cDNA microarray studies indicated that age-related defects in cellular proliferation are associated with diminished expression of the FoxM1B transcription factor. Here, we show that increased levels of FoxM1B in regenerating liver of old transgenic mice restore the sharp peaks in hepatocyte DNA replication and mitosis that are the hallmarks of young regenerating mouse liver. Restoration of the young regenerating liver phenotype is associated with increased expression of numerous cell cycle regulatory genes that include cyclin D1, cyclin A2, cyclin F, cyclin B1, cyclin B2, Cdc25B, and p55cdc. Cotransfection assays in the human hepatoma HepG2 cell line demonstrated that FoxM1B protein stimulated expression of both the cyclin B1 and cyclin D1 promoters, suggesting that these cyclin genes are a direct FoxM1B transcriptional target. These results suggest that FoxM1B controls the transcriptional network of genes that are essential for cell division and exit from mitosis. Our results indicate that reduced expression of the FoxM1B transcription factor contributes to the decline in cellular proliferation observed in the aging process.T he mammalian liver is one of the few adult organs capable of completely regenerating itself in response to injury through the release of growth factors that stimulate reentry of terminally differentiated hepatocytes into the cell cycle (1-3). Liver regeneration induced by a two-thirds partial hepatectomy (PHx) results in synchronous induction of sharp peaks in hepatocyte DNA replication (S phase) and mitosis (M phase), which requires participation of the IL-6-signaling pathway (3-5). The forkhead box (Fox) family of transcription factors (6) shares homology in the winged-helix DNA-binding domain (7), and its members play important roles in regulating transcription of genes involved in cellular proliferation, differentiation, metabolic homeostasis, longevity, and cellular transformation (8-18). The mammalian (human) Fox family member FoxM1B (previously known as HFH-11B or trident) is a ubiquitously expressed transcription factor restricted to proliferating cells of the mouse embryo (including liver) and is essential for embryonic development (19), but its expression diminishes during postnatal cellular differentiation (20). In regenerating liver, FoxM1B expression is reactivated before DNA replication (S phase) and sustained throughout the period of hepatocyte proliferation (20). Liver regeneration studies with transgenic mice, in which the transthyretin (TTR) promoter functioned to prematurely express FoxM1...

Transcriptional Activation. The liver performs essential functions in the body by uniquely expressing hepatocyte-specific genes encoding plasma proteins, clotting factors and enzymes involved in detoxification, gluconeogenesis, glycogen synthesis, and glucose, fat, and cholesterol metabolism. 1 Functional analysis of numerous hepatocyte-specific DNA regulatory regions (promoter and enhancer regions) reveals that they are composed of multiple cis-acting DNA sequences that bind different families of liver-enriched transcription factors named hepatocyte nuclear factors (HNF). The analysis involved transient transfections of hepatoma cells lines utilizing DNA regulatory regions containing targeted mutations in each of these transcription factor binding sites fused to a reporter gene, whose expression levels were used to monitor promoter activity. [2][3][4][5][6][7][8][9][10][11]

Single‐atom catalysts have drawn considerable attention because of their unique catalytic properties. However, the high surface energy of single atoms restricts their fabrication and creates significant challenges for further developments. In order to overcome this problem, metal organic framework (MOF)‐derived carbon materials can be served as ideal supports to anchor atomically dispersed metal atoms, due to their tunable particle size and shape features, by providing high surface area, porosity, thermal, and chemical stability. This review highlights the recent advances in i) different types of construction strategies for MOF‐derived carbon‐supported single‐atom catalysts, and ii) the catalytic applications of these MOF‐derived carbon‐supported single‐atom catalysts. Further, this review offers a valuable insight into the current challenges and future opportunities for MOF‐derived carbon‐supported single‐atom catalysts.