Xiao Fang Yu

Overland heroin trafficking routes and HIV-1 spread in south and south-east Asia

ObjectivesBurma produces approximately 60% of the worlds heroin, Laos is the third leading producer. Recent outbreaks of injecting drug use and HIV-1 in Burma, India, China, and Vietnam have been associated with Burmese and Laotian overland heroin trafficking routes. We analyzed findings from narcotics investigations, molecular epidemiology studies of HIV-1, and epidemiologic and behavioral studies of injecting drug use, to evaluate the roles that the heroin export routes play in the spread of drug use and HIV-1 in south and south-east Asia. MethodsWe reviewed the medical and narcotics literature, the molecular epidemiology of HIV, and did key informant interviews in India, China, and Burma with injecting drug users, drug traffickers, public health staff, and narcotics control personnel. ResultsFour recent outbreaks of HIV-1 among injecting drug users appear linked to trafficking routes. Route 1: From Burmas eastern border to Chinas Yunnan Province, with initial spread of HIV-1 subtype B, and later C. Route 2: Eastern Burma to Yunnan, going north and west, to Xinjiang Province, with B, C, and a B/C recombinant subtype. Route 3: Burma and Laos, through northern Vietnam, to Chinas Guangxi Province, subtype E. Route 4: Western Burma, across the Burma–India border to Manipur, predominant subtype C, and B and E. ConclusionsOverland heroin export routes have been associated with dual epidemics of injecting drug use and HIV infection in three Asian countries and along four routes. Molecular epidemiology is useful for mapping heroin routes. Single country narcotics and HIV programs are unlikely to succeed unless the regional narcotic-based economy is addressed.

Guidelines for Naming Nonprimate APOBEC3 Genes and Proteins

Guidelines for Naming Nonprimate APOBEC3 Genes and Proteins Rebecca S. LaRue, Valgerdur Andresdottir, Yannick Blanchard, Silvestro G. Conticello, David Derse, Michael Emerman, Warner C. Greene, Stefan R. Jonsson, Nathaniel R. Landau, Martin Lochelt, Harmit S. Malik, Michael H. Malim, Carsten Munk, Stephen J. O’Brien, Vinay K. Pathak, Klaus Strebel, Simon Wain-Hobson, Xiao-Fang Yu, Naoya Yuhki, and Reuben S. Harris*

Structural Insight into the Human Immunodeficiency Virus Vif SOCS Box and Its Role in Human E3 Ubiquitin Ligase Assembly

ABSTRACT Human immunodeficiency virus (HIV) virion infectivity factor (Vif) causes the proteasome-mediated destruction of human antiviral protein APOBEC3G by tethering it to a cellular E3 ubiquitin ligase composed of ElonginB, ElonginC, Cullin5, and Rbx2. It has been proposed that HIV Vif hijacks the E3 ligase through two regions within its C-terminal domain: a BC box region that interacts with ElonginC and a novel zinc finger motif that interacts with Cullin5. We have determined the crystal structure of the HIV Vif BC box in complex with human ElonginB and ElonginC. This complex presents direct structural evidence of the recruitment of a human ubiquitin ligase by a viral BC box protein that mimics the conserved interactions of cellular ubiquitin ligases. We further mutated conserved hydrophobic residues in a region downstream of the Vif BC box. These mutations demonstrate that this region, the Vif Cullin box, composes a third E3-ligase recruiting site critical for interaction between Vif and Cullin5. Furthermore, our homology modeling reveals that the Vif Cullin box and zinc finger motif may be positioned adjacent to the N terminus of Cullin5 for interaction with loop regions in the first cullin repeat of Cullin5.

Multimeric Soluble CD40 Ligand and GITR Ligand as Adjuvants for Human Immunodeficiency Virus DNA Vaccines

ABSTRACT For use in humans, human immunodeficiency virus (HIV) DNA vaccines may need to include immunostimulatory adjuvant molecules. CD40 ligand (CD40L), a member of the tumor necrosis factor (TNF) superfamily (TNFSF), is one candidate adjuvant, but it has been difficult to use because it is normally expressed as a trimeric membrane molecule. Soluble trimeric forms of CD40L have been produced, but in vitro data indicate that multimeric, many-trimer forms of soluble CD40L are more active. This multimerization requirement was evaluated in mice using plasmids that encoded either 1-trimer, 2-trimer, or 4-trimer soluble forms of CD40L. Fusion with the body of Acrp30 was used to produce the 2-trimer form, and fusion with the body of surfactant protein D was used to produce the 4-trimer form. Using plasmids for secreted HIV-1 antigens Gag and Env, soluble CD40L was active as an adjuvant in direct proportion to the valence of the trimers (1 < 2 < 4). These CD40L-augmented DNA vaccines elicited strong CD8+ T-cell responses but did not elicit significant CD4+ T-cell or antibody responses. To test the applicability of the multimeric fusion protein approach to other TNFSFs, a 4-trimer construct for the ligand of glucocorticoid-induced TNF family-related receptor (GITR) was also prepared. Multimeric soluble GITR ligand (GITRL) augmented the CD8+ T-cell, CD4+ T-cell, and antibody responses to DNA vaccination. In summary, multimeric CD40L and GITRL are new adjuvants for DNA vaccines. Plasmids for expressing multimeric TNFSF fusion proteins permit the rapid testing of TNFSF molecules in vivo.

Emerging HIV infections with distinct subtypes of HIV-1 infection among injection drug users from geographically separate locations in Guangxi province, China

Heroin users from Guangxi province, a southern province of China that borders Vietnam in the south and Yunnan province in China in the west, were studied for prevalence and risk factors for HIV-1 infection. Viral env sequences from HIV-1-positive individuals were also determined for subtypes of HIV-1. The overall HIV prevalence among 227 heroin users was 40%. Most had used drugs for < or = 3 years. Sharing of injection equipment and unprotected sex were significantly associated with HIV-1 infection. Subtypes C and E HIV-1 were detected in infected heroin users and were sharply segregated in two geographic locations: only subtype C was found in a border city with Yunnan province, whereas only subtype E was found in a city bordering northern Vietnam. HIV-1 strains within each subtype were remarkably homogenous, with a mean intersubject DNA distance of 2.32% for subtype E and 1.13% for subtype C, respectively. Phylogenetic analysis of C2-V5 region of Guangxi subtype E env sequences revealed significant clustering with subtype E sequences from southern Vietnam and Cambodia. These results suggest that HIV-1 infection among heroin users in Guangxi represents two emerging epidemics initiated from distinct sources: one from Vietnam and another from Yunnan province. Factors associated with HIV-1 infection were not restricted to injection practices. Unprotected sexual behaviors are likely to increase the probability of HIV transmission beyond this high-risk population. Designing and implementing effective intervention strategies targeted toward both injection drug use and high risk sexual behavior are urgently needed to further reduce HIV-1 spread in China.

Two subtypes of HIV-1 among injection-drug users in southern China

The rate of HIV-1 infection has increased steadily in China by about 80% annually and by the end of September 1997, 8277 HIV-1 cases had been reported, of whom more than 75% were IV drug users (IVDUs). UNAIDS, however, has estimated that up to 200,000 people could actually be infected with HIV-1 in China. Guangxi Province borders Yunnan province in the west and Vietnam to the south, and is a major transit area for heroin trafficking from the opium-growing region of Laos and Myanmar. Phylogenetic analyses of HIV-1 env sequences (C2-V3) obtained from 14 IVDUs found that 9 subjects from Pingxiang City were infected with subtype E and 5 from Baise City with subtype C. The 9 subtype E and 5 subtype C HIV-1 sequences were clustered together within each group, with significant bootstrap values of 100% and 95%, respectively. The subtype E sequences were more closely related to HIV-1 subtype E from Thailand than to those from Africa, and the subtype C sequences were clustered more closely to those from India than to those from Africa. Study results suggest 2 epidemiologically unrelated epidemics and 2 different sources; subtype C probably transmitted from Yunnan to Baise City through drug trafficking and IVDU interaction, and subtype E coming into China from Vietnam.

A Patch of Positively Charged Amino Acids Surrounding the Human Immunodeficiency Virus Type 1 Vif SLVx4Yx9Y Motif Influences Its Interaction with APOBEC3G

ABSTRACT The amino-terminal region of the Vif molecule in human immunodeficiency virus type 1 (HIV-1), HIV-2, and simian immunodeficiency virus (SIV) contains a conserved SLV/Ix4Yx9Y motif that was first described in 1992, but the importance of this motif for Vif function has not yet been examined. Our characterization of the amino acids surrounding this motif in HIV-1 Vif indicated that the region is critical for APOBEC3 suppression. In particular, amino acids K22, K26, Y30, and Y40 were found to be important for the Vif-induced degradation and suppression of cellular APOBEC3G (A3G). However, mutation of these residues had little effect on the Vif-mediated suppression of A3F, A3C, or A3DE, suggesting that these four residues are not important for Vif assembly with the Cul5 E3 ubiquitin ligase or protein folding in general. The LV portion of the Vif SLV/Ix4Yx9Y motif was found to be required for optimal suppression of A3F, A3C, or A3DE. Thus, the SLV/Ix4Yx9Y motif and surrounding amino acids represent an important functional domain in the Vif-mediated defense against APOBEC3. In particular, the positively charged K26 of HIV-1 Vif is invariably conserved within the SLV/Ix4Yx9Y motif of HIV/SIV Vif molecules and was the most critical residue for A3G inactivation. A patch of positively charged and hydrophilic residues (K22x3K26x3Y30x9YRHHY44) and a cluster of hydrophobic residues (V55xIPLx4-5LxΦx2YWxL72) were both involved in A3G binding and inactivation. These structural motifs in HIV-1 Vif represent attractive targets for the development of lead inhibitors to combat HIV infection.

Enhancement of Primary and Secondary Cellular Immune Responses against Human Immunodeficiency Virus Type 1 Gag by Using DNA Expression Vectors That Target Gag Antigen to the Secretory Pathway

ABSTRACT In this study, we have investigated the influence of antigen targeting after DNA vaccination upon the induction of cellular immune responses against human immunodeficiency virus type 1 (HIV-1) Gag. In addition to the standard version of HIV-1 Gag, we constructed Gag expression vectors that encode a secreted (Sc-Gag) and a cytoplasmic (Cy-Gag) Gag molecule. Although all three HIV-1 Gag expression vectors induced detectable humoral and cellular immune responses, after intramuscular injection the DNA vector encoding the Sc-Gag generated the highest primary cytotoxic T-lymphocyte (CTL) and T-helper responses. Mice immunized with one of the HIV-1 Gag DNA vectors (but not with the control vector pcDNA3.1) developed a protective immune response against infection with recombinant vaccinia virus expressing HIV-1 Gag, and this response persisted for 125 days. The magnitude of the protection correlated with the levels of Gag-specific ex vivo CTL activity and the number of CD8+ T cells producing gamma interferon. The DNA vector encoding the Sc-Gag induced higher levels of protection and greater secondary CTL responses than did the DNA vector encoding Cy-Gag.

A Single Amino Acid Difference in Human APOBEC3H Variants Determines HIV-1 Vif Sensitivity

ABSTRACT Several variants of APOBEC3H (A3H) have been identified in different human populations. Certain variants of this protein are particularly potent inhibitors of retrotransposons and retroviruses, including HIV-1. However, it is not clear whether HIV-1 Vif can recognize and suppress the antiviral activity of A3H variants, as it does with other APOBEC3 proteins. We now report that A3H_Haplotype II (HapII), a potent inhibitor of HIV-1 in the absence of Vif, can indeed be degraded by HIV-1 Vif. Vif-induced degradation of A3H_HapII was blocked by the proteasome inhibitor MG132 and a Cullin5 (Cul5) dominant negative mutant. In addition, Vif mutants that were incapable of assembly with the host E3 ligase complex factors Cul5, ElonginB, and ElonginC were also defective for A3H_HapII suppression. Although we found that Vif hijacks the same E3 ligase to degrade A3H_HapII as it does to inactivate APOBEC3G (A3G) and APOBEC3F (A3F), more Vif motifs were involved in A3H_HapII inactivation than in either A3G or A3F suppression. In contrast to A3H_HapII, A3H_Haplotype I (HapI), which differs in only three amino acids from A3H_HapII, was resistant to HIV-1 Vif-mediated degradation. We also found that residue 121 was critical for determining A3H sensitivity and binding to HIV-1 Vif.

Human Immunodeficiency Virus Type 1 Vif Is Efficiently Packaged into Virions during Productive but Not Chronic Infection

ABSTRACT Packaging of the human immunodeficiency virus type 1 Vif protein into virus particles is mediated through an interaction with viral genomic RNA and results in the association of Vif with the nucleoprotein complex. Despite the specificity of this process, calculations of the amount of Vif packaged have produced vastly different results. Here, we compared the efficiency of packaging of Vif into virions derived from acutely and chronically infected H9 cells. We found that Vif was efficiently packaged into virions from acutely infected cells (60 to 100 copies per virion), while packaging into virions from chronically infected H9 cells was near the limit of detection (four to six copies of Vif per virion). Superinfection by an exogenous Vif-defective virus did not rescue packaging of endogenous Vif expressed in the chronically infected culture. In contrast, exogenous Vif expressed by superinfection of wild-type virus was readily packaged (30 to 40 copies per virion). Biochemical analyses suggest that the differences in the relative packaging efficiencies were not due to gross differences in the steady-state distribution of Vif in chronically or acutely infected cells but are likely due to differences in the relative rates of de novo synthesis of Vif. Despite its low packaging efficiency, endogenously expressed Vif was sufficient to direct the production of viruses with almost wild-type infectivity. The results from our study provide novel insights into the biochemical properties of Vif and offer an explanation for the reported differences regarding Vif packaging.