RESUMO
The purpose of this study was to investigate the balance between transfer ribonucleic acid (tRNA) supply and demand in retrovirus-infected cells, seeking the best targets for antiretroviral therapy based on the hypothetical tRNA Inhibition Therapy (TRIT). Codon usage and tRNA gene data were retrieved from public databases. Based on logistic principles, a therapeutic score (T-score) was calculated for all sense codons, in each retrovirus-host system. Codons that are critical for viral protein translation, but not as critical for the host, have the highest T-score values. Theoretically, inactivating the cognate tRNA species should imply a severe reduction of the elongation rate during viral mRNA translation. We developed a method to predict tRNA species critical for retroviral protein synthesis. Four of the best TRIT targets in HIV-1 and HIV-2 encode Large Hydrophobic Residues (LHR), which have a central role in protein folding. One of them, codon CUA, is also a TRIT target in both HTLV-1 and HTLV-2. Therefore, a drug designed for inactivating or reducing the cytoplasmatic concentration of tRNA species with anticodon TAG could attenuate significantly both HIV and HTLV protein synthesis rates. Inversely, replacing codons ending in UA by synonymous codons should increase the expression, which is relevant for DNA vaccine design.
RESUMO
BACKGROUND: Human T-cell Leukemia Virus type 1 (HTLV-1) is the etiological agent of tropical spastic paraparesis/HTLV-associated myelopathy (HAM/TSP) that can be identified in around 0.25%-3.8% of the infected population. Disease progression can be monitored by the proviral load and may depend on genetic factors, however, it is not well understood why some HTLV-1 infected people develop the disease while others do not. The present study attempts to assess the molecular diversity of gp46 glycoprotein in HAM/TSP patients and Health Carrier (HC) individuals. METHODS: Blood samples were collected from 10 individuals, and DNA was extracted from PBMCs to measure the HTLV-1 proviral load. The gp46 coding sequences were amplified PCR, cloned and sequenced. The molecular characterization was performed using bioinformatics tools. RESULTS: The median HTLV-1 proviral load of HC (n = 5) and HAM/TSP (n = 5) patients was similar (average 316,227 copies/106 PBMCs). The gp46 molecular characterization of 146 clones (70 HC and 76 HAM/TSP) revealed an overall diversity, within HC and HAM/TSP clones, of 0.4% and 0.6%, respectively. Five frequent mutations were detected among groups (HAM/TSP and HC clone sequences). A single amino acid (aa) substitution (S35L) was exclusive for the HC group, and three gp46 substitutions (F14S, N42H, G72S) were exclusive for the HAM/TSP group. The remaining frequent mutation (V247I) was present in both groups (p = 0.0014). The in silico protein analysis revealed that the mutated alleles F14S and N42H represent more hydrophilic and flexible protein domains that are likely to be less antigenic. The Receptor Binding Domain is quite variable in the HAM/TSP group. Two other domains (aa 53-75 and 175-209) that contain multiple linear T-cell epitopes showed genetic diversity in both HAM/TSP and HC groups. Further analysis revealed 27 and 13 T-cell epitopes for class I HLA alleles and class II HLA alleles, when analyzing the entire gp46. CONCLUSIONS: The most common gp46 mutations were not associated clinical status because they were found in only one individual, except for the V247I mutation, that was found at viral clones from HAM/TSP ad HC individuals. Because of this, we cannot associate any of the gp46 found mutations with the clinical profile.
Assuntos
Portador Sadio/virologia , Produtos do Gene env/genética , Vírus Linfotrópico T Tipo 1 Humano/genética , Paraparesia Espástica Tropical/virologia , Proteínas Oncogênicas de Retroviridae/genética , Adulto , Idoso , Sequência de Aminoácidos , Portador Sadio/imunologia , Epitopos de Linfócito T/química , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Produtos do Gene env/química , Produtos do Gene env/imunologia , Vírus Linfotrópico T Tipo 1 Humano/química , Vírus Linfotrópico T Tipo 1 Humano/imunologia , Vírus Linfotrópico T Tipo 1 Humano/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutação , Paraparesia Espástica Tropical/imunologia , Estrutura Terciária de Proteína , Proteínas Oncogênicas de Retroviridae/química , Proteínas Oncogênicas de Retroviridae/imunologiaRESUMO
HIV-1 provirus activation is under control of the long terminal repeat (LTR)-5' viral promoter region, which presents remarkable genetic variation among HIV-1 subtypes. It is possible that molecular features of the LTR contribute to the unusual profile of the subtype C epidemic in the Brazilian Southern region. To characterize the LTR of Brazilian HIV isolates, we analyzed sequences from 21 infected individuals from Porto Alegre and Salvador cities. Sequences were compared with subtype B and C reference strains from different countries. Phylogenetic analysis showed that 17 (81%) samples were subtype B and four (19%) were subtype C. Common patterns of transcription factor binding sites (TFBS) in subtypes B and C sequences were confirmed and other potential TFBS specific for subtype C were found. Brazilian subtype C sequences contained an additional NF-κB biding site, as previously described for the majority of subtype C isolates. The high level of LTR polymorphisms identified in this study might be important for viral fitness.
