RESUMO
For over two decades, research on Human immunodeficiency virus (HIV), which is responsible for AIDS, has aimed at understanding of the molecular mechanisms used by this virus during its life cycle. An essential step in the HIV life cycle is the budding, which promotes the release of viral particles from the host cell. It has recently been revealed that HIV in the process of budding uses besides one viral protein also the machinery of the infected cell, in particular the proteins Tsg101 and ubiquitin. The viral protein is the p6 domain of the Gag precursor polyprotein. In normal cells, Tsg101 functions as a regulator of endocytic trafficking that recognizes ubiquitinated cargo and directs its delivery to degradative compartments. In HIV-infected cells, Tsg101 and ubiquitin interact with Gag p6 to promote the release of new viral particles from the host cell. Molecular mechanisms underlying the process of HIV budding from infected cells suggests a whole new range of drug targets that could prove useful in AIDS suppression in HIV-positive patients.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Produtos do Gene gag/fisiologia , HIV/fisiologia , Fatores de Transcrição/fisiologia , Ubiquitina/fisiologia , Animais , Complexos Endossomais de Distribuição Requeridos para Transporte , HumanosRESUMO
BACKGROUND: Nef is an accessory protein of primate lentiviruses, HIV-1, HIV-2 and SIV. Besides removing CD4 and MHC class I from the surface and activating cellular signaling cascades, Nef also binds GagPol during late stages of the viral replicative cycle. In this report, we investigated further the ability of Nef to facilitate the replication of HIV-1. RESULTS: To this end, first the release of new viral particles was much lower in the absence of Nef in a T cell line. Since the same results were obtained in the absence of the viral envelope using pseudo-typed viruses, this phenomenon was independent of CD4 and enhanced infectivity. Next, we found that Nef not only possesses a consensus motif for but also binds AIP1 in vitro and in vivo. AIP1 is the critical intermediate in the formation of multivesicular bodies (MVBs), which play an important role in the budding and release of viruses from infected cells. Indeed, Nef proliferated MVBs in cells, but only when its AIP1-binding site was intact. Finally, these functions of Nef were reproduced in primary macrophages, where the wild type but not mutant Nef proteins led to increased release of new viral particles from infected cells. CONCLUSION: We conclude that by binding GagPol and AIP1, Nef not only proliferates MVBs but also contributes to the egress of viral particles from infected cells.
Assuntos
Produtos do Gene nef/metabolismo , HIV-1/metabolismo , Proteínas/metabolismo , Vesículas Transportadoras/virologia , Proteínas Adaptadoras de Transdução de Sinal , Sequência de Aminoácidos , Sítios de Ligação , Antígenos CD4/metabolismo , Proteínas de Transporte , Linhagem Celular Transformada , Sequência Consenso , Produtos do Gene gag/química , Produtos do Gene gag/fisiologia , Produtos do Gene nef/química , Produtos do Gene nef/fisiologia , Guanilato Quinases , HIV-1/patogenicidade , HIV-1/fisiologia , Humanos , Macrófagos/virologia , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Alinhamento de Sequência , Vesículas Transportadoras/metabolismo , Vesículas Transportadoras/ultraestrutura , Replicação Viral , Produtos do Gene nef do Vírus da Imunodeficiência HumanaRESUMO
Recombination is an important way to generate genetic diversity. Accumulation of HIV-1 full-length genomes in databases demonstrated that recombination is pervasive in viral strains collected globally. Recombinant forms achieving epidemiological relevance are termed circulating recombinant forms (CRFs). CRF12_BF was up to now the only CRF described in South America. The objective was to identify the first CRF in Brazil conducting full genome analysis of samples sharing the same partial genome recombinant structure. Ten samples obtained from individuals residing in Santos, Brazil, sharing the same recombination pattern based on partial genome sequence data, were selected from a larger group to undergo full length genome analysis. Near full length genomes were assembled from overlapping fragments. Mosaic genomes were evaluated by Bootscan, alignment inspection, and phylogenetic analysis using neighbor joining and maximum likelihood. Full genomes were also analyzed by split decomposition. We were able to identify five mosaic genomes. Two of these structures were represented by at least three samples derived from epidemiologically unlinked individuals. These structures were named CRF28_BF and CRF29_BF and are the second and third CRFs composed exclusively by subtypes B and F as well as the second and third CRFs encountered in South America. Other recombinant forms studied here resembled CRF28_BF and CRF29_BF. Our results suggest that a diverse population of related recombinants, including CRFs may play an important part in the Brazilian and South American epidemic.
Assuntos
Variação Genética , Infecções por HIV/virologia , HIV-1/genética , Recombinação Genética , Adulto , Brasil/epidemiologia , Produtos do Gene gag/genética , Produtos do Gene gag/fisiologia , Infecções por HIV/epidemiologia , HIV-1/isolamento & purificação , Humanos , Masculino , Epidemiologia Molecular , FilogeniaRESUMO
The findings that BF intersubtype recombinant human immunodeficiency type 1 viruses (HIV-1) with coincident breakpoints in pol are circulating widely in Argentina and that non-recombinant F subtype viruses have failed to be detected in this country were reported recently. To analyse the mosaic structures of these viruses and to determine their phylogenetic relationship, near full-length proviral genomes of eight of these recombinant viruses were amplified by PCR and sequenced. Intersubtype breakpoints were analysed by bootscanning and examining the signature nucleotides. Phylogenetic relationships were determined with neighbour-joining trees. Five viruses, each with predominantly subtype F genomes, exhibited mosaic structures that were highly similar. Two intersubtype breakpoints were shared by all viruses and seven by the majority. Of the consensus breakpoints, all nine were present in two viruses, which exhibited identical recombinant structures, and four to eight breakpoints were present in the remaining viruses. Phylogenetic analysis of partial sequences supported both a common ancestry, at least in part of their genomes, for all recombinant viruses and the phylogenetic relationship of F subtype segments with F subtype viruses from Brazil. A common ancestry of the recombinants was supported also by the presence of shared signature amino acids and nucleotides, either unreported or highly unusual in F and B subtype viruses. These results indicate that HIV-1 BF recombinant viruses with diverse mosaic structures, including a circulating recombinant form (which are widespread in Argentina) derive from a common recombinant ancestor and that F subtype segments of these recombinants are related phylogenetically to the F subtype viruses from Brazil.