Gag Sequences Research Articles

Genotypic and functional impact of HIV-1 adaptation to its host population during the North American epidemic.

HLA-restricted immune escape mutations that persist following HIV transmission could gradually spread through the viral population, thereby compromising host antiviral immunity as the epidemic progresses. To assess the extent and phenotypic impact of this phenomenon in an immunogenetically diverse population, we genotypically and functionally compared linked HLA and HIV (Gag/Nef) sequences from 358 historic (1979–1989) and 382 modern (2000–2011) specimens from four key cities in the North American epidemic (New York, Boston, San Francisco, Vancouver). Inferred HIV phylogenies were star-like, with approximately two-fold greater mean pairwise distances in modern versus historic sequences. The reconstructed epidemic ancestral (founder) HIV sequence was essentially identical to the North American subtype B consensus. Consistent with gradual diversification of a “consensus-like” founder virus, the median “background” frequencies of individual HLA-associated polymorphisms in HIV (in individuals lacking the restricting HLA[s]) were ∼2-fold higher in modern versus historic HIV sequences, though these remained notably low overall (e.g. in Gag, medians were 3.7% in the 2000s versus 2.0% in the 1980s). HIV polymorphisms exhibiting the greatest relative spread were those restricted by protective HLAs. Despite these increases, when HIV sequences were analyzed as a whole, their total average burden of polymorphisms that were “pre-adapted” to the average host HLA profile was only ∼2% greater in modern versus historic eras. Furthermore, HLA-associated polymorphisms identified in historic HIV sequences were consistent with those detectable today, with none identified that could explain the few HIV codons where the inferred epidemic ancestor differed from the modern consensus. Results are therefore consistent with slow HIV adaptation to HLA, but at a rate unlikely to yield imminent negative implications for cellular immunity, at least in North America. Intriguingly, temporal changes in protein activity of patient-derived Nef (though not Gag) sequences were observed, suggesting functional implications of population-level HIV evolution on certain viral proteins.

HIV-1 Protease-Substrate Coevolution in Nelfinavir Resistance

Resistance to various human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs) challenges the effectiveness of therapies in treating HIV-1-infected individuals and AIDS patients. The virus accumulates mutations within the protease (PR) that render the PIs less potent. Occasionally, Gag sequences also coevolve with mutations at PR cleavage sites contributing to drug resistance. In this study, we investigated the structural basis of coevolution of the p1-p6 cleavage site with the nelfinavir (NFV) resistance D30N/N88D protease mutations by determining crystal structures of wild-type and NFV-resistant HIV-1 protease in complex with p1-p6 substrate peptide variants with L449F and/or S451N. Alterations of residue 30's interaction with the substrate are compensated by the coevolving L449F and S451N cleavage site mutations. This interdependency in the PR-p1-p6 interactions enhances intermolecular contacts and reinforces the overall fit of the substrate within the substrate envelope, likely enabling coevolution to sustain substrate recognition and cleavage in the presence of PR resistance mutations. Resistance to human immunodeficiency virus type 1 (HIV-1) protease inhibitors challenges the effectiveness of therapies in treating HIV-1-infected individuals and AIDS patients. Mutations in HIV-1 protease selected under the pressure of protease inhibitors render the inhibitors less potent. Occasionally, Gag sequences also mutate and coevolve with protease, contributing to maintenance of viral fitness and to drug resistance. In this study, we investigated the structural basis of coevolution at the Gag p1-p6 cleavage site with the nelfinavir (NFV) resistance D30N/N88D protease mutations. Our structural analysis reveals the interdependency of protease-substrate interactions and how coevolution may restore substrate recognition and cleavage in the presence of protease drug resistance mutations.

Molecular epidemiology related to human immunodeficiency virus type 1 infection in men having sex with men in Xi'an

To investigate the subtype distribution of HIV-1 infection among men having sex with men (MSM) in Xi'an city. 5 ml anti-coagulating blood samples were collected from MSM who had been reported during 2010-2012 in Xi'an. Both gag and env genes were amplified by nested RT-PCR from the extracted RNA and then sequenced. The acquired sequences were compared with international subtype references, and then the genetic distances were calculated and phylogenetic trees were constructed by Mega 5.2. Epidemiological information including sexual behavior characteristics, history of drug use, blood donation, etc. were gathered. 168 samples were successfully amplified and sequenced. Results from Phylogenic analysis showed that the gag and env sequences of 165 samples shared the same subtypes, of which 79 (47.0%) were CRF01_AE, 74 (44.0%) were CRF07_BC and 12 (7.1%) belonged to subtype B. There were 3 samples with gag and env sequences classified into different subtypes, of which 2(1.2%) were CRF01_AE/A1, and the other 1 was CRF07_BC/CRF01_AE. At least three HIV-1 subtypes including CRF01_AE, CRF07_BC were identified among MSM population in Xi'an city.

Jaagsiekte sheep retrovirus detected in human lung cancer tissue arrays

BackgroundAdenocarcinoma is the most common type of non-small cell lung cancer and is frequently observed in non-smoking patients. Adenocarcinoma in-situ (formerly referred to as bronchioloalveolar carcinoma) is a subset of lung adenocarcinoma characterized by growth along alveolar septae without evidence of stromal, vascular, or pleural invasion, that disproportionately affects never-smokers, women, and Asians. Adenocarcinoma in-situ is morphologically and histologically similar to a contagious lung neoplasm of sheep called ovine pulmonary adenocarcinoma (OPA). OPA is caused by infection with the exogenous betaretrovirus, jaagsiekte sheep retrovirus (JSRV), whose envelope protein (Env) is a potent oncogene. Several studies have reported that a proportion of human lung adenocarcinomas are immunopositive for an antigen related to the Gag protein of JSRV, however other groups have been unable to verify these observations by PCR.MethodsHere we examine human lung cancer tissue arrays (TA) for evidence of JSRV Env protein and DNA by immunohistochemical staining and PCR, respectively.ResultsOur results reveal that a subset of human lung cancers express an antigen that reacts with a JSRV Env-specific monoclonal antibody in immunohistochemistry and that exogenous JSRV-like env and gag sequences can be amplified from TA tumor samples, albeit inefficiently.ConclusionsWhile a causative role has not been established, these data suggest that a JSRV-like virus might infect humans. With next generation sequencing approaches, a JSRV-like virus in human lung cancers may be identified which could have profound implications for prevention, diagnosis and therapy.

Tos17 rice element: incomplete but effective

BackgroundTos17 was the first LTR retrotransposon (Copia) described as active in cultivated rice, and is present in two copies in the genome of the sequenced Nipponbare variety. Only the chromosome 7 copy is active and able to retrotranspose, at least during in vitro culture, and this ability was widely used in insertional mutagenesis assays.ResultsHere the structure of the active Tos17 was thoroughly annotated using a set of bioinformatic analyses.ConclusionsUnexpectedly, Tos17 appears to be a non-autonomous LTR retrotransposon, lacking the gag sequence and thus unable to transpose by itself.

HIV-1 Replication Fitness of HLA-B*57/58:01 CTL Escape Variants Is Restored by the Accumulation of Compensatory Mutations in Gag

Expression of HLA-B*57 and the closely related HLA-B*58:01 are associated with prolonged survival after HIV-1 infection. However, large differences in disease course are observed among HLA-B*57/58:01 patients. Escape mutations in CTL epitopes restricted by these HLA alleles come at a fitness cost and particularly the T242N mutation in the TW10 CTL epitope in Gag has been demonstrated to decrease the viral replication capacity. Additional mutations within or flanking this CTL epitope can partially restore replication fitness of CTL escape variants. Five HLA-B*57/58:01 progressors and 5 HLA-B*57/58:01 long-term nonprogressors (LTNPs) were followed longitudinally and we studied which compensatory mutations were involved in the restoration of the viral fitness of variants that escaped from HLA-B*57/58:01-restricted CTL pressure. The Sequence Harmony algorithm was used to detect homology in amino acid composition by comparing longitudinal Gag sequences obtained from HIV-1 patients positive and negative for HLA-B*57/58:01 and from HLA-B*57/58:01 progressors and LTNPs. Although virus isolates from HLA-B*57/58:01 individuals contained multiple CTL escape mutations, these escape mutations were not associated with disease progression. In sequences from HLA-B*57/58:01 progressors, 5 additional mutations in Gag were observed: S126N, L215T, H219Q, M228I and N252H. The combination of these mutations restored the replication fitness of CTL escape HIV-1 variants. Furthermore, we observed a positive correlation between the number of escape and compensatory mutations in Gag and the replication fitness of biological HIV-1 variants isolated from HLA-B*57/58:01 patients, suggesting that the replication fitness of HLA-B*57/58:01 escape variants is restored by accumulation of compensatory mutations.

Study on Correlation of Maedi-Visna Virus (MVV) with Ovine Subclinical Mastitis in Iran.

Maedi-Visna is an important slow viral disease of sheep leading to progressive pneumonia, encephalitis and mastitis. Udder is one of the organs affected by MVV. Despite the fact that in Iran Maedi-Visna is known since 2000, to the authors' knowledge correlation of subclinical mastitis and infection with MVV has not been assayed. In this study 50 milk samples from 10 flocks in East Azerbaijan Province of Iran were tested. None of the animals exhibited any clinical signs of the disease. Forty samples were collected from CMT positive ewes and ten were taken from CMT negative ewes. Milk samples were analyzed using PCR targeting gag sequence. Presence of provirus DNA was detected in one sample from CMT negative and seven samples from CMT positive ewes. These data demonstrate that 16.5% of sheep with subclinical mastitis were infected to MVV. Thus this virus can be considered one of the main pathogenic agents of mastitis and can be dramatically transmitted to lambs by milk.

Increased Sequence Coverage through Combined Targeting of Variant and Conserved Epitopes Correlates with Control of HIV Replication

A major challenge in the development of an HIV vaccine is that of contending with the extensive sequence variability found in circulating viruses. Induction of HIV-specific T-cell responses targeting conserved regions and induction of HIV-specific T-cell responses recognizing a high number of epitope variants have both been proposed as strategies to overcome this challenge. We addressed the ability of cytotoxic T lymphocytes from 30 untreated HIV-infected subjects with and without control of virus replication to recognize all clade B Gag sequence variants encoded by at least 5% of the sequences in the Los Alamos National Laboratory HIV database (1,300 peptides) using gamma interferon and interleukin-2 (IFN-γ/IL-2) FluoroSpot analysis. While targeting of conserved regions was similar in the two groups (P = 0.47), we found that subjects with control of virus replication demonstrated marginally lower recognition of Gag epitope variants than subjects with normal progression (P = 0.05). In viremic controllers and progressors, we found variant recognition to be associated with viral load (r = 0.62, P = 0.001). Interestingly, we show that increased overall sequence coverage, defined as the overall proportion of HIV database sequences targeted through the Gag-specific repertoire, is inversely associated with viral load (r = -0.38, P = 0.03). Furthermore, we found that sequence coverage, but not variant recognition, correlated with increased recognition of a panel of clade B HIV founder viruses (r = 0.50, P = 0.004). We propose sequence coverage by HIV Gag-specific immune responses as a possible correlate of protection that may contribute to control of virus replication. Additionally, sequence coverage serves as a valuable measure by which to evaluate the protective potential of future vaccination strategies.

Molecular Dating of HIV-1 Subtype C from Bangladesh

Bangladesh has an overall low HIV prevalence of <0.1% in the general population and <1% among key affected populations, but it is one of few Asian countries that has yet to reverse the epidemic. In order to do this, it is important to understand the transmission dynamics in this country. The aim of this study was to investigate the phylogenetic relationships of HIV-1 subtype C strains from Bangladesh and related strains from other countries, and thereby clarify when and from where subtype C was introduced in the country and how it subsequently spread within Bangladesh. The phylogenetic analysis included 118 Bangladeshi gag sequences and 128 sequences from other countries and was performed using the BEAST package. Our analysis revealed that the vast majority of Bangladeshi sequences (97/118, 82%) fall into a large regional cluster of samples from Bangladesh, India, China and Myanmar, which dates back to the early 1960’s. Following its establishment in the region, this strain has entered Bangladesh multiple times from around 1975 and onwards, but extensive in-country transmission could only be detected among drug users and not through sexual transmission. In addition, there have been multiple (at least ten) introductions of subtype C to Bangladesh from outside this region, but no extensive spread could be detected for any of these. Since many HIV-infections remain undetected while asymptomatic, the true extent of the transmission of each strain remains unknown, especially among hard to reach groups such as clients of sex workers and returning migrants with families.

Functional conservation of HIV-1 Gag: implications for rational drug design

BackgroundHIV-1 replication can be successfully blocked by targeting gag gene products, offering a promising strategy for new drug classes that complement current HIV-1 treatment options. However, naturally occurring polymorphisms at drug binding sites can severely compromise HIV-1 susceptibility to gag inhibitors in clinical and experimental studies. Therefore, a comprehensive understanding of gag natural diversity is needed.FindingsWe analyzed the degree of functional conservation in 10862 full-length gag sequences across 8 major HIV-1 subtypes and identified the impact of natural variation on known drug binding positions targeted by more than 20 gag inhibitors published to date. Complete conservation across all subtypes was detected in 147 (29%) out of 500 gag positions, with the highest level of conservation observed in capsid protein. Almost half (41%) of the 136 known drug binding positions were completely conserved, but all inhibitors were confronted with naturally occurring polymorphisms in their binding sites, some of which correlated with HIV-1 subtype. Integration of sequence and structural information revealed one drug binding pocket with minimal genetic variability, which is situated at the N-terminal domain of the capsid protein.ConclusionsThis first large-scale analysis of full-length HIV-1 gag provided a detailed mapping of natural diversity across major subtypes and highlighted the considerable variation in current drug binding sites. Our results contribute to the optimization of gag inhibitors in rational drug design, given that drug binding sites should ideally be conserved across all HIV-1 subtypes.

SHAPE analysis of the 5′ end of the Mason-Pfizer monkey virus (MPMV) genomic RNA reveals structural elements required for genome dimerization

Earlier genetic and structural prediction analyses revealed that the packaging determinants of Mason Pfizer monkey virus (MPMV) include two discontinuous core regions at the 5' end of its genomic RNA. RNA secondary structure predictions suggested that these packaging determinants fold into several stem-loops (SLs). To experimentally validate this structural model, we employed selective 2' hydroxyl acylation analyzed by primer extension (SHAPE), which examines the flexibility of the RNA backbone at each nucleotide position. Our SHAPE data validated several predicted structural motifs, including U5/Gag long-range interactions (LRIs), a stretch of single-stranded purine (ssPurine)-rich region, and a distinctive G-C-rich palindromic (pal) SL. Minimum free-energy structure predictions, phylogenetic, and in silico modeling analyses of different MPMV strains revealed that the U5 and gag sequences involved in the LRIs differ minimally within strains and maintain a very high degree of complementarity. Since the pal SL forms a helix loop containing a canonical "GC" dyad, it may act as a RNA dimerization initiation site (DIS), enabling the virus to package two copies of its genome. Analyses of wild-type and pal mutant RNAs revealed that disruption of pal sequence strongly affected RNA dimerization. However, when in vitro transcribed trans-complementary pal mutants were incubated together showed RNA dimerization was restored authenticating that the pal loop (5'-CGGCCG-3') functions as DIS.

Small ruminant lentivirus genetic subgroups associate with sheep TMEM154 genotypes

Small ruminant lentiviruses (SRLVs) are prevalent in North American sheep and a major cause of production losses for the U.S. sheep industry. Sheep susceptibility to SRLV infection is influenced by genetic variation within the ovine transmembrane 154 gene (TMEM154). Animals with either of two distinct TMEM154 haplotypes that both encode glutamate at position 35 of the protein (E35) are at greater risk of SRLV infection than those homozygous with a lysine (K35) haplotype. Prior to this study, it was unknown if TMEM154 associations with infection are influenced by SRLV genetic subgroups. Accordingly, our goals were to characterize SRLVs naturally infecting sheep from a diverse U.S. Midwestern flock and test them for associations with TMEM154 E35K genotypes. Two regions of the SRLV genome were targeted for proviral amplification, cloning, sequence analysis, and association testing with TMEM154 E35K genotypes: gag and the transmembrane region of env. Independent analyses of gag and env sequences showed that they clustered in two subgroups (1 and 2), they were distinct from SRLV subtypes originating from Europe, and that subgroup 1 associated with hemizygous and homozygous TMEM154 K35 genotypes and subgroup 2 with hemi- and homozygous E35 genotypes (gag p < 0.001, env p = 0.01). These results indicate that SRLVs in the U.S. have adapted to infect sheep with specific TMEM154 E35K genotypes. Consequently, both host and SRLV genotypes affect the relative risk of SRLV infection in sheep.

Phylogenetic analysis of small ruminant lentiviruses in mixed flocks: Multiple evidence of dual infection and natural transmission of types A2 and B1 between sheep and goats

Previous molecular analyses of small ruminant lentivirus (SRLV) populations in single species herds in Quebec, Canada, have revealed a relatively simple structure where goats and sheep appeared exclusively infected with B1 and A2 subtypes respectively. The present work aimed at extending these earlier findings with the analysis of SRLVs in mixed flocks. Molecular analyses revealed a more complex picture of SRLV population structure in mixed herds compared to single species herds. Notably, phylogenetic analyses of long gag sequences strongly support transmission of A2 subtype from sheep to goats as well as transmission of B1 subtype from goats to sheep. Hence, this work uncovered for the first time natural transmission between sheep and goats of North American subtype A2. In addition, multiple evidences of mixed infection of sheep and goats with A2 and B1 subtypes were found. The data reported in this study reinforces the concept of a genetic continuum of SRLVs where strains are exchanged between sheep and goats under favourable conditions and in the absence of specific species barriers. Most interestingly, this study suggests that dual infection, which is a hallmark of the lentivirus paradigm HIV, may not be such rare events in small ruminants but may simply be understudied and underreported. Overall, the present data shows that sheep and goats in Canada can be infected with both SRLV A and B types, sometimes simultaneously, and that mixed flocks may represent a breeding ground for their evolution.

Anti-retroviral drugs used to treat a patient with chronic lymphocytic leukemia and cancer-related fatigue.

e22039 Background: The only known human cancer retrovirus, HTLV1 causes adult T-cell leukemia/lymphoma of which some respond to AZT+ alpha interferon. The existence of human retroviruses originating from murine leukemia viruses (MLVs) and their role in human cancer has been debated since the 1970s. A previously untreated 68 y.o. Oncologist with B-cell CLL and Cancer-Related Fatigue (CRF) was studied for retrovirus to determine if he could be treated with anti-retroviral drugs (arvs). His history included autoimmune disorders. Methods: Antibody detection: Alter et al. 2012. Gag detection: Mikovits et al. 2010. Results: Antibody to MLV envelope antigen and Gag RNA was detected. The Gag sequence was similar to MLVs but sufficiently different from the laboratory generated, XMRV which has no natural history in Man to rule it out. Proinflammatory cytokines were elevated. Trisomy 12 was present. By day 571 of his CLL the ALC doubling time decreased to &lt; 1 year and he developed disabling CRF. AZT and raltegravir were started day 899. ALC, cytokine levels and CRF improved. After 9 months CLL parameters relapsed and then responded to adding tenofovir. This response lasted 3 months and then relapsed. Addition of lopinavir induced a response of parameters now ongoing 9 months. Total time on treatment is 29 months without toxicity and with improved CRF. In addition to his CLL there was a measurable T-cell clone and monocytosis both of which rose and fell with relapses and responses of the CLL to arvs but at different rates. Conclusions: A common human neoplasm was successfully treated with arvs. The drugs used were 2 Reverse Transcriptase inhibitors, an Integrase inhibitor and a Protease inhibitor developed for HIV treatment. There is nothing unusual or unique about this patient’s neoplasm and his case should stimulate further research. The role of retrovirus in cancer and CRF and whether arvs could be beneficial to patients identified as having a retroviral associated cancer and CRF should be studied. In addition, the significance of this patient’s clonal T-cell expansion and monocytosis and their production of cytokines should be examined. [Table: see text]

Prevalence of Transmitted HIV Drug Resistance in Iran between 2010 and 2011

ObjectiveDrug-resistant (DR) HIV emerges during combined antiretroviral treatment (cART), creating concern about widespread transmission of DR-HIV as cART is expanded in resource-limited countries. The aim of this study was to determine the predominant HIV-1 subtypes and prevalence of transmitted DR mutations among antiretroviral-naïve patients in Iran.DesignTo monitor transmission of DR HIV, a threshold surveillance based on the world health organization (WHO) guidelines was implemented in Iran.MethodsFor this HIVDR threshold surveillance study, blood samples were collected from 50 antiretroviral-naïve HIV-1-infected patients. Antiretroviral-resistant mutations were determined by sequencing HIV-1 protease, reverse transcriptase and integrase regions. The HIV-1 subtype was determined by sequencing the p17 and C2-V5 regions of the gag and env genes, respectively.ResultsPhylogenetic analyses of the sequenced regions revealed that 45 (95.7%) of 47 samples that were successfully obtained were CRF35_AD. The remaining two cases were subtype B (2.1%) and CRF01_AE (2.1%). Consistent results were obtained also from Env and Gag sequences. Regarding prevalence of transmitted DR viruses, two cases were found to harbor reverse transcriptase-inhibitor-resistant mutations (4.3%). In addition, although not in the WHO list for surveillance of transmitted mutations, 13 minor protease-inhibitor-resistant mutations listed in the International AIDS Society-USA panel of drug resistance mutations were found. No DR mutations were detected in the integrase region.ConclusionsOur study clarified that CRF35_AD is the major subtype among HIV-1-infected patients in Iran. According to the WHO categorization method of HIVDR threshold survey, the prevalence of transmitted drug resistant HIV in Iran was estimated as moderate (5–15%).

Genetic characterization of small ruminant lentiviruses circulating in naturally infected sheep and goats in Ontario, Canada

Maedi–visna virus (MVV) and caprine arthritis encephalitis virus (CAEV) are related members of a group of small ruminant lentiviruses (SRLVs) that infect sheep and goats. SRLVs are endemic in many countries, including Canada. However, very little is known about the genetic characteristics of Canadian SRLVs, particularly in the province of Ontario. Given the importance of surveillance and eradication programs for the control of SRLVs, it is imperative that the diagnostic tests used to identify infected animals are sensitive to local strains of SRLVs. The aim of this work was to characterize SRLV strains circulating in Ontario and to evaluate the variability of the immunodominant regions of the Gag protein. In this study, the nearly complete gag sequence of 164 SRLVs, from 130 naturally infected sheep and 32 naturally infected goats from Ontario, was sequenced. Animals belonged to distantly located single and mixed species (sheep and goats) farms. Ovine lentiviruses from the same farm tended to cluster more closely together than did caprine lentiviruses from the same farm. Sequence analysis revealed a higher degree of heterogeneity among the caprine lentivirus sequences with an average inter-farm pairwise DNA distance of 10% and only 5% in the ovine lentivirus group. Interestingly, amplification of SRLVs from ELISA positive sheep was successful in 81% of cases, whereas amplification of SRLV proviral DNA was only possible in 55% of the ELISA positive goat samples; suggesting that a significant portion of caprine lentiviruses circulating in Ontario possess heterogeneity at the primer binding sites used in this study. Sequences of sheep and goat SRLVs from Ontario were assembled into phylogenetic trees with other known SRLVs and were found to belong to sequence groups A2 and B1, respectively, as defined by Shah et al. (2004a). A novel caprine lentivirus with a pairwise genetic difference of 15.6–25.4% relative to other group B subtypes was identified. Thus we suggest the designation of a novel subtype, B4, within the caprine lentivirus-like cluster. Lastly, we demonstrate evidence of recombination between ovine lentiviruses. These results emphasize the broad genetic diversity of SRLV strains circulating in the province of Ontario and show that the gag region is suitable for phylogenetic studies and may be applied to monitor SRLV eradication programs.

Foamy Virus Assembly with Emphasis on Pol Encapsidation

Foamy viruses (FVs) differ from all other genera of retroviruses (orthoretroviruses) in many aspects of viral replication. In this review, we discuss FV assembly, with special emphasis on Pol incorporation. FV assembly takes place intracellularly, near the pericentriolar region, at a site similar to that used by betaretroviruses. The regions of Gag, Pol and genomic RNA required for viral assembly are described. In contrast to orthoretroviral Pol, which is synthesized as a Gag-Pol fusion protein and packaged through Gag-Gag interactions, FV Pol is synthesized from a spliced mRNA lacking all Gag sequences. Thus, encapsidation of FV Pol requires a different mechanism. We detail how WT Pol lacking Gag sequences is incorporated into virus particles. In addition, a mutant in which Pol is expressed as an orthoretroviral-like Gag-Pol fusion protein is discussed. We also discuss temporal regulation of the protease, reverse transcriptase and integrase activities of WT FV Pol.

Study on the association between genetic variation of gag gene and HLA restricted CTL response in HIV-1 B' infected Chinese people

To identify the genetic variation of Gag gene in HIV-1 B' subtype infected Chinese people and to evaluate the association between the positive selection sites and HLA restricted CTL response. We downloaded all HIV-1 B' subtype gag gene sampled from China from the LANL HIV database. The sequences were grouped by years of collection and the mean intra-subtype diversity and divergence were examined by MEGA 5.0 software. The sites under positive selection were estimated by using Hyphy software. Then analyzed the association between positively selected sites and HLA restricted CTL epitopes. A set of 122 B' subtype HIV-1 gag sequences were collected from Henan, Hubei, Liaoning and Yunnan provinces, grouped by sampling year as < 2001, 2001 - 2003, 2004 - 2006 and 2007 - 2010. The mean intra-subtype diversity and divergence increased over time (r(1) = 0.497, P1 < 0.001; r(2) = 0.593, P2 < 0.001). This analysis revealed 34 positively selected sites and 97.1% of these sites were located within 47 known CTL epitopes. The most frequent HLA was HLA-A*02 (21.6%) followed by A*11(9.8%), B*35(9.8%) and B*57(5.9%). The percentages of HLA-A/B/C limited CTL epitopes were 43.1%, 49.0% and 7.9%, respectively. The mean intra-subtype diversity and divergence of B' subtype gag sequences increased over time. B' subtype HIV-1 virus has accumulated adaptations to HLA restricted CTL responses in HIV-1 infected Chinese.

A cis-Acting Element in Retroviral Genomic RNA Links Gag-Pol Ribosomal Frameshifting to Selective Viral RNA Encapsidation

During retroviral RNA encapsidation, two full-length genomic (g) RNAs are selectively incorporated into assembling virions. Packaging involves a cis-acting packaging element (Ψ) within the 5' untranslated region of unspliced HIV-1 RNA genome. However, the mechanism(s) that selects and limits gRNAs for packaging remains uncertain. Using a dual complementation system involving bipartite HIV-1 gRNA, we observed that gRNA packaging is additionally dependent on a cis-acting RNA element, the genomic RNA packaging enhancer (GRPE), found within the gag p1-p6 domain and overlapping the Gag-Pol ribosomal frameshift signal. Deleting or disrupting the two conserved GRPE stem loops diminished gRNA packaging and infectivity >50-fold, while deleting gag sequences between Ψ and GRPE had no effect. Downregulating the translation termination factor eRF1 produces defective virus particles containing 20 times more gRNA. Thus, only the HIV-1 RNAs employed for Gag-Pol translation may be specifically selected for encapsidation, possibly explaining the limitation of two gRNAs per virion.

Genetic Characterization and Transmitted Drug Resistance of the HIV Type 1 Epidemic in Men Who Have Sex with Men in Beijing, China

A rapid increase in the number of HIV cases in the men who have sex with men (MSM) population has been observed in China; however, little information is available on the genetic characterization of HIV prevalent in this population. In this study, 95 HIV-1-seropositive drug-naive patients from the Beijing MSM population were enrolled. The genetic characterization and transmission of drug resistance of HIV-1 were examined based on full-length gag, pol, and partial env gene sequences. Three subtypes, including CRF01_AE (56.0%), B (30.8%), and CRF07_BC (12.6%), were identified. Close phylogenetic relationships were found among these strains with isolates from other populations in Beijing and MSM isolates from Hebei province, which suggested that the Beijing MSM population might act as a bridge for HIV transmission between MSM and other high-risk populations. Drug-resistant mutations were identified in 5.3% of sampled individuals. Our results provided detailed genetic data and would be helpful for understanding the transmitting pattern of HIV strains between MSM and other populations.