Transcriptional activation of gammaherpesviral oncogene promoters by the hepatitis B viral X protein (HBx)

The latent membrane protein-1 (LMP1) of Epstein-Barr Virus (EBV), saimiri transformation protein (STP) of Herpesvirus saimiri (HVS), and K1 protein of Kaposi's sarcoma-associated herpesvirus (KSHV) are potent gammaherpesvirus oncogenes. To study the possible effects of double viral infection, we investigated the effects of oncogenic early proteins of DNA viruses E1A and E1B (adenovirus-5), E6 and E7 (human papillomavirus-16), HBx (hepatitis B virus), Tag (SV40), and gammaherpesviral oncogene during co-infection in human B-lymphoma (Ramos) and human T-cell leukemia (Jurkat) cell lines. HBx transactivated the promoters of LMP1, STP, and K1 the most, by about six-, three-, and twofold, respectively. Analyses of site-directed mutation and the heterologous promoter system showed that HBx activated the promoter activity of these genes via the NF-kappaB site. These results suggest that HBV (HBx) infection of cells previously infected by gammaherpesviruses transactivates their oncogenes, resulting in possible virus-related disease pathogenesis.     

Neutrophils activation can be diminished by apolipoprotein A-I

High density lipoprotein (HDL) has anti-inflammatory function. To investigate the effects of apolipoprotein A-I (ApoA-I), the major apolipoprotein of HDL, on activated neutrophils, we stimulated neutrophils in vitro with fMLP and PMA, as a receptor-binding and a nonreceptor-binding stimuli, respectively, and incubated ApoA-I with those neutrophils. Three conditions were utilized: 1) resting neutrophils + ApoA-I (0, 2.5,5, 10 microg/mL respectively), 2) fMLP(10(-7) mol/L)-activated neutrophils + ApoA-I (0, 2.5, 5, 10 microg/mL respectively), and 3) PMA(10(-7) mol/L)-activated neutrophils + ApoA-I (0, 2.5, 5, 10 microg/mL respectively). After incubation, we measured neutrophils adhesion to fibronectin, oxidative bust (O2- and H2O2 production), degranulation (release of MPO and elastase), and L929 cell mortality which were attacked by release-out of cytokines in activated neutrophils (using MTT). Our results showed that in vitro ApoA-I inhibits fMLP- and PMA- activated neutrophil adhesion, oxidative burst, degranulation and L929 cell mortality. These inhibition effects of ApoA-I on fMLP-activated neutrophils are more powerful than that on PMA-activated neutrophils. ApoA-I has no effect on resting neutrophils. We concluded that ApoA-I could diminish the function of activated neutrophils.     

An RNA-binding domain in the thyroid hormone receptor enhances transcriptional activation

Thyroid hormone plays important roles in development, differentiation, and metabolic homeostasis by binding to nuclear thyroid hormone receptors, which regulate target gene expression by interacting with DNA response elements and coregulatory proteins. We show that thyroid hormone receptors also are single-stranded RNA binding proteins and that this binding is functionally significant. By using a series of deletion mutants, a novel RNA-binding domain was localized to a 41-amino acid segment of thyroid hormone receptor alpha1 between the second zinc finger and the ligand-binding domain. This RNA-binding domain was necessary and sufficient for thyroid hormone receptor binding to the steroid receptor RNA activator (SRA). Although SRA does not bind directly to steroid receptors, it has been identified as a steroid receptor coactivator, and was thought not to be a coactivator for thyroid hormone receptors. However, transfection studies revealed that SRA enhances thyroid hormone induction of appropriate reporter genes and that the thyroid hormone receptor RNA-binding domain is important for this enhancement. We conclude that thyroid hormone receptors bind RNA through a novel domain and that the interaction of this domain with SRA, and perhaps other RNAs, enhances thyroid hormone receptor function.     

Exposure on cell surface and extensive arginine methylation of ewing sarcoma (EWS) protein

In contrast to the knowledge regarding the function of chimeric Ewing sarcoma (EWS) fusion proteins that arise from chromosomal translocation, the cellular function of the RNA binding EWS protein is poorly characterized. EWS protein had been found mainly in the nucleus. In this report we show that EWS protein is not only found in the nucleus and cytosol but also on cell surfaces. After cell-surface biotinylation, isoelectric focusing of membrane fraction, avidin-agarose extraction of biotinylated proteins, and SDS-polyacrylamide gel electrophoresis, EWS protein was identified by matrix-assisted laser desorption ionization and nanoelectrospray tandem mass spectrometry of in-gel-digested peptides. These analyses revealed that the protein, having repeated RGG motifs, is extensively asymmetrically dimethylated on arginine residues, the sites of which have been mapped by mass spectrometric methods. Out of a total of 30 Arg-Gly sequences, 29 arginines were found to be at least partially methylated. The Arg-Gly-Gly sequence was present in 21 of the 29 methylation sites, and in contrast to other methylated proteins, only 11 (38%) methylated arginine residues were found in the Gly-Arg-Gly sequence. The presence of Gly on the C-terminal side of the arginine residue seems to be a prerequisite for recognition by a protein-arginine N-methyltransferase (PRMT) catalyzing this asymmetric dimethylation reaction. One monomethylarginine and no symmetrically methylated arginine residue was found. The present findings imply that RNA-binding EWS protein shuttles from the nucleus to the cell surface in a methylated form, the role of which is discussed.