Retroviral recombination as an adjunct to viral propagation and pathogenesis.
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Biomedical subjects
Publications and source records attributed to R Desrosiers.
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The nef gene is important for the pathogenicity associated with simian immunodeficiency virus infection in rhesus monkeys and with human immunodeficiency virus type 1 (HIV-1) infection in humans. The mechanisms by which nef contributes to pathogenesis in vivo remain unclear. We investigated the contribution of nef to HIV-1 replication in human lymphoid tissue ex vivo by studying infection with parental HIV-1 strain NL4-3 and with a nef mutant (DeltanefNL4-3). In human tonsillar histocultures, NL4-3 replicated to higher levels than DeltanefNL4-3 did. Increased virus production with NL4-3 infection was associated with increased numbers of productively infected cells and greater loss of CD4(+) T cells over time. While the numbers of productively infected T cells were increased in the presence of nef, the levels of viral expression and production per infected T cell were similar whether the nef gene was present or not. Exogenous interleukin-2 (IL-2) increased HIV-1 production in NL4-3-infected tissue in a dose-dependent manner. In contrast, DeltanefNL4-3 production was enhanced only marginally by IL-2. Thus, Nef can facilitate HIV-1 replication in human lymphoid tissue ex vivo by increasing the numbers of productively infected cells and by increasing the responsiveness to IL-2 stimulation.
In recent years, Actinobacillus suis, Haemophilus parasuis, and Streptococcus suis have emerged as important pathogens of swine, particularly in high health status herds. Their association with a wide range of serious clinical conditions and has given rise to the moniker "suis-ide diseases." These organisms are early colonizers and, for that reason, are difficult to control by management procedures such as segregated early weaning. Vaccination, serodiagnostic testing, and even serotyping are complicated by the presence of multiple serotypes, cross-reactive antigens, and the absence of clear markers for virulence. In this review, we discuss our current understanding of the pathogenesis, epidemiology, and management of the causative agents of the "suis-ide diseases" of swine.
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Sera from 154 African non-human primates were screened for the presence of antibodies to type D retrovirus proteins. Four of five talapoin monkeys (Miopithecus sp.) captured in western Africa were positive for antibodies to type D retrovirus by ELISA and by immunoblot reactivity. Talapoins are the only African non-human primates that have so far shown evidence for type D retrovirus infection. Thus, talapoin monkeys appear to be a reservoir of type D retrovirus infection.
The relationship between gene expression and the patterns of histone methylation in Drosophila cells has been investigated using inhibitors of transcription acting at various levels. Inhibition of ribosomal RNA synthesis and processing by 5-fluorouridine or of general RNA synthesis by camptothecin, an inhibitor of topoisomerase I, does not affect the methylation pattern of core histones. This suggests that the arrest of transcription per se is not involved in the changes in histone methylation such as those encountered in heat-shocked cells. However, ethidium bromide and novobiocin, which are known to disrupt nucleosome structure, and VM-26 (teniposide), a specific inhibitor of topoisomerase II, induce changes in histone methylation patterns which, though less severe, are similar to those observed under cellular stress. These results suggest that chromatin conformation is probably an important factor in the accessibility of histones to methyltransferases.
Heat shock or arsenite treatment alter the pattern of histone methylation in Drosophila cells. Both types of stress induce a rapid increase in the methylation level of histone H2B. The methylated amino acid residue of H2B has been identified by thin layer chromatography and electrophoresis as methylproline and is located at the N-terminal end of H2B. Heat shock also induces a decrease in the level of methylation of histone H3. Under normal growth temperature conditions, histone H3 is shown to be methylated on lysine residues. However under heat shock conditions, there is a decrease in the extent of methylation of lysine residues and the appearance of new methylation on arginine residues in H3. These new heat shock-induced methylated residues have been identified as the symmetrical and asymmetrical forms of dimethylarginine. The methylated amino acid residue of histone H4 is lysine with mono-, di-, and trimethyl forms found in both control and heat or chemically stressed cells. These stress-induced changes in the methylation level of the N-terminal proline residue of histone H2B and shift in the methylation sites of histone H3 may be involved in the restructuration of chromatin accompanying the inactivation of normal genes in response to stress. Moreover, we suggest that the hypermethylation of H2B may also be involved in its protection from increased ubiquitin-mediated proteolytic activity under these conditions of cellular stress.
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Biochemical and immunological properties of retrovirus-D/New England (here referred as R-D/NE) recently isolated at the New England Regional Primate Research Center, Southborough, Ma, from a rhesus monkey with acquired immune deficiency syndrome were investigated and compared to the prototype type D retroviruses Mason-Pfizer monkey virus (MPMV) and permanent human fibroblast virus (PMFV) isolated from a breast carcinoma of a rhesus monkey and a continuous human cell line, respectively. The polypeptide composition of R-D/NE propagated in a human lymphoid B cell line (Raji cells) has been investigated using SDS-polyacrylamide gel electrophoresis. Staining with Coomassie blue and labelling with 14C amino acids revealed seven viral polypeptides with molecular weights of 4,000, 10,000, 12,000, 15,000, 18,000, 27,000, and 80,000 Da which were also shared by MPMV and PMFV. The 80,000 Da protein was shown to be a glycoprotein by incorporation of 3H glucosamine. The 18,000 Da protein was identified as a phosphoprotein of R-D/NE. p18 structural proteins of MPMV and PMFV represent phosphoproteins of their respective viruses as well. All three phosphorylated proteins contain O-phosphoserine as major phosphoamino acid. The comparison of tryptic peptide maps of the major internal structural proteins of R-D/NE, MPMV, and PMFV revealed a striking similarity among p 10/p 12 and p 15. proteins. A minor difference was detected among the tryptic peptide digests of p 4 and p 27 proteins. Antiserum against p 15 of MPMV showed a significantly weaker binding to R-D/NE than to MPMV and PMFV at high dilutions.
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The effects of a heat shock or arsenite treatment on the methylation and acetylation of core histones have been investigated in Drosophila cultured cells. The decrease in H3 methylation, which is observed during a heat shock, is not a demethylation process, but results from methylation arrest. Two-dimensional gel electrophoresis leaves no ambiguity concerning the identity of H2B as a methylated protein, since H2B and D2, a nuclear nonhistone protein, which comigrate on one-dimensional gels, are well separated on these gels. Two-dimensional gel electrophoresis in the presence of Triton X-100 resolves each of the core histones into multiple forms resulting from posttranslational modifications. There are apparently, however, no histone variants in cultured Drosophila cells. At 23 degrees C, the various forms of the core histones resolved on two-dimensional gels are methylated. Under heat-shock or arsenite treatment, the methylation of all forms of H3 is decreased, while that of the various forms of H2B increase. These stress conditions also induce a generalized diminution in the acetylation of all forms of core histones. In the course of a heat shock, the synthesis of H2B is increased and this newly synthesized histone remains unacetylated during the shock. These changes in the patterns of core histone methylation and acetylation may be correlated with the reorganization of gene activity brought about by the heat shock.
The potential significance of the important post-synthetic methylation changes of core histones observed upon heat shock or after an arsenite treatment has been studied in Drosophila Kc cells. The changes in the histone methylation patterns appeared to be linked with the repression of normal gene expression rather than with the activation of the heat shock genes. Thus following a brief heat shock, the kinetics of recovery of the normal pattern of gene expression could be correlated with those of the return to a normal pattern of methylation of histones H2B and H3. Experiments with different inducers and a transcriptional inhibitor also suggest that the decreased methylation of H3 and the increase in that of H2B are an effect of the repression of transcription processes.
Anesthetic management during tracheobronchial reconstruction is a concern to the anesthetist, who must maintain satisfactory gas exchange while ensuring adequate exposure to the trachea. The technique for high-flow catheter ventilation was first described for bronchoscopy, and it involves positive-pressure breathing with a high flow (40 to 60 L/min) of oxygen. This flow is directed to a semirigid catheter inserted in the endotracheal tube and around which the tracheobronchial anastomosis can be done without interruption. The value of the technique was tested in 18 patients undergoing tracheobronchial reconstructions. Patients' ages ranged from 22 to 69 years and the average duration of catheter ventilation was 35 minutes. Regardless of the duration of high-flow catheter ventilation good arterial blood gas values were maintained in all patients. In six patients, the average oxygen tension (measured at 5 minute intervals) was 416 mm Hg and the average carbon dioxide tension was 34 mm Hg. One patient developed surgical emphysema during the procedure. The high-flow catheter ventilation provides specific advantages during tracheobronchial procedures: avoidance of endotracheal manipulations, unobstructed field during surgical reconstruction, and good oxygenation throughout the procedure.
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Extracts from interferon-treated, not virus infected EAT cells differ in several biochemical characteristics from extracts of untreated cells. Some of these differences are manifested only if the extracts are supplemented with ds RNA and ATP. Thus, in the extracts from interferon-treated cells these supplements activate a protein kinase and an endonuclease activity as well as an inhibitor of the translation of messenger RNA. The effect of the same supplements in extracts of untreated cells is much less pronounced. Other differences between the two types of extracts do not seem to depend on the addition of ds RNA and ATP. These include an impairment of mRNA cap methylation and an inhibition of peptide chain elongation that can be overcome by the addition of tRNA. The treatment of human (HeLa S3) cells with human interferon is manifested in the cell extract similarly to the treatment of EAT cells with mouse interferon. Studies are underway to isolate and characterize the ds RNA activated enzymes and the inhibitors and to establish how the presence of these in extracts from interferon-treated cells can account for the impairment of virus replication by interferon.