RESUMEN
Nef proteins from all primate Lentiviruses, including the simian immunodeficiency virus of chimpanzees (SIVcpz), increase viral progeny infectivity. However, the function of Nef involved with the increase in viral infectivity is still not completely understood. Nonetheless, until now, studies investigating the functions of Nef from SIVcpz have been conducted in the context of the HIV-1 proviruses. In an attempt to investigate the role played by Nef during the replication cycle of an SIVcpz, a Nef-defective derivative was obtained from the SIVcpzWTGab2 clone by introducing a frame shift mutation at a unique restriction site within the nef sequence. This nef-deleted clone expresses an N-terminal 74-amino acid truncated peptide of Nef and was named SIVcpz-tNef. We found that the SIVcpz-tNef does not behave as a classic nef-deleted HIV-1 or simian immunodeficiency virus of macaques SIVmac. Markedly, SIVcpz-tNef progeny from both Hek-293T and Molt producer cells were completely non-infectious. Moreover, the loss in infectivity of SIVcpz-tNef correlated with the inhibition of Gag and GagPol processing. A marked accumulation of Gag and very low levels of reverse transcriptase were detected in viral lysates. Furthermore, these observations were reproduced once the tNef peptide was expressed in trans both in SIVcpzΔNef and HIV-1WT expressing cells, demonstrating that the truncated peptide is a dominant negative for viral processing and infectivity for both SIVcpz and HIV-1. We demonstrated that the truncated Nef peptide binds to GagPol outside the protease region and by doing so probably blocks processing of both GagPol and Gag precursors at a very early stage. This study demonstrates for the first time that naturally-occurring Nef peptides can potently block lentiviral processing and infectivity.
Asunto(s)
Productos del Gen nef/metabolismo , VIH-1/fisiología , Virus de la Inmunodeficiencia de los Simios/fisiología , Replicación Viral , Animales , Línea Celular , Mutación del Sistema de Lectura , Técnicas de Inactivación de Genes , Productos del Gen gag/metabolismo , Productos del Gen nef/genética , Productos del Gen pol/metabolismo , Humanos , Pan troglodytes , Unión Proteica , Virus de la Inmunodeficiencia de los Simios/genéticaRESUMEN
BACKGROUND: Nucleoside/nucleotide analogue (NA) treatment causes selection pressure for HBV strains carrying mutations conferring NA resistance. Drug-resistance mutations occur in the reverse transcriptase (RT) region of the HBV polymerase gene and spontaneously arise during viral replication. These mutations can also alter the hepatitis B surface (HBs) protein and in some cases reduce binding to HBs antibodies. The spread of NA-resistant HBV may impact the efficacy of antiviral treatment and hepatitis B immunization programmes. In this study, we used direct sequencing to assess the occurrence of HBV carrying known mutations that confer NA resistance in the largest cohort of treatment-naive patients with chronic hepatitis B (CHB) to date. METHODS: HBV DNA samples isolated from 702 patients were sequenced and the RT region subjected to mutational analysis. RESULTS: There was high genetic variability among the HBV samples analysed: A1 (63.7%), D3 (14.5%), A2 (3.3%), A3 (0.1%), B1 (0.1%), B2 (0.1%), C2 (0.9%), D1 (0.9%), D2 (4.6%), D4 (5.1%), D unclassified subgenotype (0.7%), E (0.6%), F2a (4.6%), F4 (0.4%) and G (0.4%). HBV strains harbouring mutations conferring NA resistance alone or combined with compensatory mutations were identified in 1.6% (11/702) of the patients. CONCLUSIONS: HBV strains harbouring resistance mutations can comprise the major population of HBV quasispecies in treatment-naive patients. In Brazil, there is a very low frequency of untreated patients who are infected with these strains. These findings suggest that the spread and natural selection of drug-resistant HBV is an uncommon event and/or most of these strains remain unstable in the absence of NA selective pressure.
Asunto(s)
Antivirales/farmacología , Farmacorresistencia Viral/genética , Productos del Gen pol/genética , Virus de la Hepatitis B/efectos de los fármacos , Hepatitis B Crónica/virología , Mutación , Adenina/análogos & derivados , Adenina/farmacología , Anticuerpos Antivirales/genética , Anticuerpos Antivirales/inmunología , Brasil , ADN Viral/genética , ADN Viral/inmunología , Productos del Gen pol/antagonistas & inhibidores , Productos del Gen pol/metabolismo , Genotipo , Guanina/análogos & derivados , Guanina/farmacología , Antígenos de Superficie de la Hepatitis B/genética , Antígenos de Superficie de la Hepatitis B/inmunología , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/inmunología , Hepatitis B Crónica/diagnóstico , Hepatitis B Crónica/inmunología , Humanos , Lamivudine/farmacología , Pruebas de Sensibilidad Microbiana , Organofosfonatos/farmacología , Estudios Retrospectivos , Análisis de Secuencia de ADN , Tenofovir/farmacología , Replicación Viral/efectos de los fármacosRESUMEN
Feline immunodeficiency virus (FIV) is the Lentivirus responsible for an immunodeficiency-like disease in domestic cats (Felis catus). FIV is divided into five phylogenetic subtypes (A, B, C, D, and E), based on genetic diversity. Knowledge of the geographical distribution of subtypes is relevant for understanding different disease progressions and for vaccine development. In this study, viral sequences of 26 infected cats from Rio de Janeiro, 8 undergoing treatment with zidovudine (AZT) for at least 5 years, were successfully amplified from blood specimens. gag capsid (CA), pol reverse transcriptase (RT), and env gp120 (V3-V4) regions were analyzed to determine subtypes and to evaluate potential mutations related to antiretroviral drug resistance among treated cats. Subtyping based on phylogenetic analysis was performed by the neighbor-joining and maximum likelihood methods. All of the sequences clustered with subtype B in the three regions, exhibiting low genetic variability. Additionally, we found evidence that the same virus is circulating in animals in close contact. The analysis of FIV RT sequences identified two new putative mutations related to drug resistance located in the RT "finger" domain, which has 60% identity to human immunodeficiency virus (HIV) sequence. Amino acid change K-->R at codons 64 and 69 was found in 25% and 37.5% of the treated animals, respectively. These signatures were comparable to K65R and K70R thymidine-associated mutations found in the HIV-1 HXB2 counterpart. This finding strongly suggests a position correlation between the mutations found in FIV and the K65R and K70R substitutions from drug-resistant HIV-1 strains.