Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ability of minigene-encoded viral peptide epitopes to be presented by class I molecules in the absence of MHC-encoded transporters has been evaluated in mutant T2 cells. These cells have a large deletion in the class II MHC region that includes the known transporter protein for antigenic peptides and proteasome genes and they are defective in presenting viral epitopes to CTL. T2 cells that express minigenes encoding the influenza virus matrix peptide 58-66 (GILGFVFTL) and two HTLV 1 Tax peptides 11-19 (LLFGYPVYV) and 12-19 were lysed by HLA-A2-restricted peptide-specific CTL. Minigene expression of a HLA-A2-restricted HIV reverse transcriptase peptide 476-484 (ILKEPVHGV) with three charged residues sensitized T2 cells poorly for lysis by HIV-specific CTL unless the peptide was preceded by an endoplasmic reticulum translocation signal sequence. Expression of an influenza virus nucleoprotein peptide 383-391 (SRYWAIRTR) with three charged arginine residues did sensitize HLA-B27+ T2 cells for lysis by peptide-specific CTL. These and other results with endogenously expressed peptide analogs in which hydrophobic and charged amino acids were interchanged demonstrate that antigenic peptides can be translocated from the cytoplasm into the class I Ag presentation pathway independent of MHC-encoded transporters; and that peptide hydrophobicity appears not to be a major determinant in selecting peptides for this alternate pathway.
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PMID:Presentation of endogenous peptides to MHC class I-restricted cytotoxic T lymphocytes in transport deletion mutant T2 cells. 767 94

The class II region of the mammalian MHC harbors two proteasome subunit genes, LMP2 and LMP7. These genes are induced by IFN-gamma, and their products are incorporated into proteasomes substituting for their closest relatives, the delta and X subunits, respectively. This substitution is believed to change the proteolytic specificity of proteasomes, making it more suitable for generation of peptides to be presented by class I molecules. To elucidate the phylogenetic origin of LMP2 and the linkage of its gene with the MHC, reverse transcriptase-PCR amplification of Xenopus laevis and lamprey liver mRNA was performed with primers designed to amplify both the mammalian LMP2 and delta sequences. Both LMP2 and delta were amplified from X. laevis, whereas only delta was amplified from lamprey, suggesting that delta/LMP2 gene duplication occurred after divergence of cyclostomes but before divergence of amphibians. The linkage between the LMP2 gene and the MHC was observed in a diploid Xenopus species, Xenopus tropicalis, but not in a tetraploid species, X. laevis, indicating that this linkage was established before the divergence of amphibian from higher vertebrates, but that this linkage was lost in X. laevis, probably by a gene reorganization accompanying the tetraploidization. The X. laevis LMP2 and LMP7 mRNA showed a similar tissue distribution, indicating that the genetic linkage is not required for apparently coordinated tissue-specific expression of these genes. Sequence and linkage analyses suggest that LMP2 may not play as vital a role as LMP7 in Ag presentation.
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PMID:Evolution of proteasome subunits delta and LMP2: complementary DNA cloning and linkage analysis with MHC in lower vertebrates. 921 89

The hepatic stellate cell (HSC), following a fibrogenic stimulus, is transformed from a quiescent to an activated cell. Cytokines induce NFkappaB activity in activated but not in quiescent HSCs with subsequent expression of NFkappaB-responsive genes, such as intercellular adhesion molecule (ICAM)-1 and interleukin (IL)-6. We investigated the effect of proteasome inhibitors and an IkappaB super-repressor on the cytokine mediated activation of NFkappaB, ICAM-1, and IL-6 in activated HSCs. Culture-activated HSCs were stimulated with IL-1beta or tumor necrosis factor alpha (TNFalpha) in the presence or absence of proteasome inhibitors, ALLN or MG-132, or after infection with an adenovirus expressing the IkappaB super-repressor (Ad5IkappaB) or beta-galactosidase (Ad5LacZ) as a control. NFkappaB activity was evaluated by immunofluorescence and by electrophoretic mobility shift assay. The steady state level of cytoplasmic IkappaB protein was measured by Western Blot. ICAM-1 and IL-6 expression was measured by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbant assay. Proteasome inhibitors, which block the degradation of IkappaB, and the Ad5IkappaB, which provides an exogenous nondegradable IkappaB, block the stimulation of NFkappaB activity by TNFalpha and IL-1beta in activated HSCs. These reagents block the subsequent nuclear translocation of p65 NFkappaB and induction of ICAM-1 and IL-6 by cytokines. The specificities of the proteasome inhibitors and the IkappaB super-repressor are demonstrated by their failure to block c-Jun N-terminal kinase induction by cytokines. Cytokine-induced stimulation of NFkappaB, ICAM-1, and IL-6 is blocked by proteasome inhibitors and Ad5IkappaB in activated HSCs. Inhibition of IkappaBalpha degradation is a potential target for anti-inflammatory therapy in the liver and might influence the activation process of HSCs following fibrotic stimuli.
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PMID:Inhibition of NFkappaB in activated rat hepatic stellate cells by proteasome inhibitors and an IkappaB super-repressor. 958 6

The proteasome, an essential component of the ATP-dependent proteolytic pathway in eukaryotic cells, is responsible for the degradation of most cellular proteins and is believed to be the main source of MHC class I-restricted antigenic peptides for presentation to CTL. Inhibition of the proteasome by lactacystin or various peptide aldehydes can result in defective Ag presentation, and the pivotal role of the proteasome in Ag processing has become generally accepted. However, recent reports have challenged this observation. Here we examine the processing requirements of two HLA A*0201-restricted epitopes from HIV-1 reverse transcriptase and find that they are produced by different degradation pathways. Presentation of the C-terminal ILKEPVHGV epitope is impaired in ME275 melanoma cells by treatment with lactacystin, and is independent of expression of the IFN-gamma-inducible proteasome beta subunits LMP2 and LMP7. In contrast, both lactacystin treatment and expression of LMP7 induce the presentation of the N-terminal VIYQYMDDL epitope. Consistent with these observations we show that up-regulation of LMP7 by IFN-gamma enhances presentation of the VIYQYMDDL epitope. Hence interplay between constitutive and IFN-gamma-inducible beta-subunits of the proteasome can qualitatively influence Ag presentation. These observations may have relevance to the patterns of immunodominance during the natural course of viral infection.
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PMID:IFN-gamma exposes a cryptic cytotoxic T lymphocyte epitope in HIV-1 reverse transcriptase. 1035 50

The activated 20 S proteasome, which has been found only in mammalian cells, is composed of two heptamer rings of an activator protein on each end of the 20 S proteasome and is inducible by interferon-gamma. A 20 S proteasome has been recently identified in a protozoan pathogen Trypanosoma brucei, but there has been no experimental evidence yet for the presence of a 26 S proteasome. Instead, an activated form of 20 S proteasome was isolated from this organism, which has significantly enhanced peptidase activities. It consists of an additional activator protein with an estimated molecular mass of 26 kDa (PA26) (To, W. Y., and Wang, C. C. (1997) FEBS Lett. 404, 253-262). The profile and sequences of tryptic peptides from PA26 were determined by mass spectrometry; no matches were found in the data base. The peptide sequences were used in reverse transcriptase-polymerase chain reaction to isolate a full-length cDNA clone encoding PA26. The protein sequence thus derived from it indicates little sequence identity with those of mammalian activator proteins PA28 alpha, beta, or gamma. There is only a single copy of PA26 gene in T. brucei. Purified recombinant PA26 polymerizes spontaneously to form heptamer ring with an outer diameter of 8.5 nm. The ring binds and activates 20 S proteasomes from T. brucei as well as rat, whereas human PA28alpha can neither bind nor activate T. brucei 20 S proteasome. The former is thus apparently more ubiquitous than PA28 in its capability of binding to and activating 20 S proteasomes. Its presence in T. brucei may also suggest a more ancient origin of proteasome activator proteins and a much wider involvement in protein degradation among other eukaryotic organisms than was originally envisaged.
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PMID:Structural and functional characterizations of the proteasome-activating protein PA26 from Trypanosoma brucei. 1056 54

A means of regulating the fate of intracellular proteins is their covalent conjugation to ubiquitin-like proteins. A recently discovered ubiquitin-like protein is called "diubiquitin" because it consists of two ubiquitin-like domains in head-to-tail arrangement. Human diubiquitin is encoded at the telomeric end of the MHC class I locus and was previously found to be expressed in dendritic cells and mature B cells. We have extended the expression analysis of diubiquitin by reverse transcriptase-PCR and Northern blotting in primary endothelial cells and human cancer cell lines derived from nine different tissues. Diubiquitin expression was found to be generally and synergistically inducible with the cytokines IFN-gamma and TNF-alpha but not with IFN-alpha. Diubiquitin mRNA expression was induced within 2 h after cytokine stimulation and was independent of protein neosynthesis but dependent on proteasome activity. The mouse homologue of diubiquitin which is also encoded in the MHC class I locus was likewise induced with IFN-gamma and TNF-alpha. A general and synergistic induction with IFN-gamma and TNF-alpha suggests that diubiquitin may exert its functions in antigen presentation or other cellular processes controlled by these two cytokines.
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PMID:A ubiquitin-like protein which is synergistically inducible by interferon-gamma and tumor necrosis factor-alpha. 1060 13

The influence of the gene expression of critical components of the cytoplasmic and lysosomal proteolytic pathways on the rate of protein degradation was evaluated in the leg skeletal muscle of 8 severely traumatized patients. Muscle proteolysis was determined as the intramuscular phenylalanine rate of appearance by L-[ring-2H5]phenylalanine infusion and the leg arteriovenous catheterization technique combined with muscle biopsy. Muscle mRNA levels of UbB polyubiquitin and cathepsin B were determined by reverse transcriptase-competitive polymerase chain reaction and expressed as a percent of the mRNA level of the housekeeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In the patients, individual values for UbB polyubiquitin mRNA levels directly correlated with the rate of muscle proteolysis (r = .76, P < .05), whereas no correlation (r = .10) was found between cathepsin B mRNA levels and proteolysis. Thus, after trauma, the rate of muscle proteolysis appears to be largely regulated by the ubiquitin-proteasome system at the level of gene transcription.
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PMID:Contribution of the ubiquitin-proteasome pathway to overall muscle proteolysis in hypercatabolic patients. 1087 90

The mutant catalase purified previously from acatalasemic dog liver was heat-labile but possessed normal activity, suggesting a mutation within the coding region distal from the catalytic site. The nucleotide and deduced amino acid sequences of acatalasemic beagle dog catalase were determined by analysis of cDNA obtained by 5'- and 3'-RACE and reverse transcriptase-polymerase chain reaction (RT-PCR) methods. Comparative analysis of cDNA sequences of normal and acatalasemic dog catalases indicated a single nucleotide difference where alanine(327) (G macro CT) was substituted with threonine (ACT). The mutant catalase, which was overexpressed in COS-1 cells, was heat-labile as previously observed with the purified enzyme from acatalasemic dog liver, indicating that this amino acid substitution can lead to structural instability. No catalase protein and activity were detected by immunoblotting and spectrophotomeric assay in acatalasemic dog reticulocytes although almost the same level of mRNA expression as that in the normal reticulocytes was observed. Pulse-labeling and immunoprecipitation examination indicated that the level of catalase synthesis in the acatalasemic dog reticulocytes was almost the same (approximately 80%) as that in the normal reticulocytes. On the other hand, the synthesized mutant catalase in reticulocytes was rapidly degraded (t(1/2): 1.8 h) compared with the normal catalase (t(1/2): 14.0 h) and this degradation was almost completely inhibited by lactacystin (LC). These results suggested that the proteolytic degradation mediated most likely by proteasome might be involved in disposing of the mutant catalase in acatalasemic erythroid cells.
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PMID:cDNA cloning of mutant catalase in acatalasemic beagle dog: single nucleotide substitution leading to thermal-instability and enhanced proteolysis of mutant enzyme. 1113 58

Widespread utilization of highly active antiretroviral therapy (HAART) for HIV-infection, primarily protease inhibitors in combination with nucleoside analogue reverse transcriptase inhibitors, has recently led to a sustained reduction in the morbidity and mortality of this disease. However, administration of HAART is frequently associated with the development of lipid disorders. The severity and prevalence of dyslipidaemia vary, depending on the type of HAART, nutritional status, HIV disease stage, and concomitant presence of lipodystrophy and insulin resistance (two additional adverse effects of HAART). The mechanism that is responsible for HAART-associated dyslipidaemia remains incompletely understood. Recent data indicate that this effect may be, at least in part, accounted for by protease inhibitor-mediated inhibition of the proteasome activity and accumulation of the active portion of sterol regulatory element-binding protein-1c in liver cells and adipocytes. Whether lipid disorders in HIV-infected patients receiving HAART translate into an increased cardiovascular risk, and the indications for lipid-lowering interventions in this population, remain to be established.
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PMID:Antiretroviral therapy-associated hyperlipidaemia in HIV disease. 1135 35

Human immunodeficiency virus type-1 (HIV-1) reverse transcriptase (RT) plays a central role in the virus replication cycle. We found that HIV-1 RT was rapidly degraded when incubated with cell extracts obtained from human peripheral blood cells. The proteolytic activity responsible for the in vitro degradation of RT was present in monocytes and their precursors. Interestingly, this activity was downregulated upon cell activation or differentiation along the macrophage pathway. The proteolytic process appears specific for HIV-1 RT since other HIV-1 proteins were not degraded upon incubation in the same extracts. Although the degradation of RT was unaffected by specific proteasome inhibitors, it could be inhibited by PMSF and aprotinin, suggesting the involvement of a serine protease. Upon cell fractionation, this serine protease was found to be associated with the microsomal fraction and displayed an apparent molecular weight of approximately 2000 kDa, as determined by gel filtration. Our results suggest that a giant serine protease, different from tripeptidyl peptidase II, is involved in the in vitro degradation of HIV-1 RT. The possibility of an in vivo interaction between HIV-1 RT and a cell-type-specific serine protease is discussed.
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PMID:Human monocytes possess a serine protease activity capable of degrading HIV-1 reverse transcriptase in vitro. 1146 30


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