To determine the protective potential of the humoral immune response against HIV-1 in vivo we evaluated the potency of three neutralizing antibodies (2G12, 2F5 and 4E10) in suppressing viral rebound in six acutely and eight chronically HIV-1-infected individuals undergoing interruption of antiretroviral treatment (ART). Only two of eight chronically infected individuals showed evidence of a delay in viral rebound during the passive immunization. Rebound in antibody-treated acutely infected individuals upon cessation of ART was substantially later than in a control group of 12 individuals with acute infection. Escape mutant analysis showed that the activity of 2G12 was crucial for the in vivo effect of the neutralizing antibody cocktail. By providing further direct evidence of the potency, breadth and titers of neutralizing antibodies that are required for in vivo activity, these data underline both the potential and the limits of humoral immunity in controlling HIV-1 infection.

Next-generation sequencing (NGS) technologies enable new insights into the diversity of virus populations within their hosts. Diversity estimation is currently restricted to single-nucleotide variants or to local fragments of no more than a few hundred nucleotides defined by the length of sequence reads. To study complex heterogeneous virus populations comprehensively, novel methods are required that allow for complete reconstruction of the individual viral haplotypes. Here, we show that assembly of whole viral genomes of ∼8600 nucleotides length is feasible from mixtures of heterogeneous HIV-1 strains derived from defined combinations of cloned virus strains and from clinical samples of an HIV-1 superinfected individual. Haplotype reconstruction was achieved using optimized experimental protocols and computational methods for amplification, sequencing and assembly. We comparatively assessed the performance of the three NGS platforms 454 Life Sciences/Roche, Illumina and Pacific Biosciences for this task. Our results prove and delineate the feasibility of NGS-based full-length viral haplotype reconstruction and provide new tools for studying evolution and pathogenesis of viruses.

The CC-chemokine RANTES (regulated on activation normal T-cell expressed and secreted; CCL5) transduces multiple intracellular signals. Like all chemokines, it stimulates G protein-coupled receptor (GPCR) activity through interaction with its cognate chemokine receptor(s), but in addition also activates a GPCR-independent signaling pathway. Here, we show that the latter pathway is mediated by an interaction between RANTES and glycosaminoglycan chains of CD44. We provide evidence that this association, at both low, physiologically relevant, and higher, probably supraphysiologic concentrations of RANTES, induces the formation of a signaling complex composed of CD44, src kinases, and adapter molecules. This triggers the activation of the p44/42 mitogen-activated protein kinase (MAPK) pathway. By specifically reducing CD44 expression using RNA interference we were able to demonstrate that the p44/p42 MAPK activation by RANTES requires a high level of CD44 expression. As well as potently inhibiting the entry of CCR5 using HIV-1 strains, RANTES can enhance HIV-1 infectivity under certain experimental conditions. This enhancement process depends in part on the activation of p44/p42 MAPK. Here we show that silencing of CD44 in HeLa-CD4 cells prevents the activation of p44/p42 MAPK and leads to a substantial reduction in HIV IntroductionThe chemokine RANTES (regulated on activation normal T-cell expressed and secreted; CCL5) is a member of the CC-chemokine family, a group of small proteins with a highly conserved tertiary. 1,2 Chemokines recruit and activate specific leukocyte populations. Multiple chemokine receptors with partially overlapping specificities for ligand binding have been described, suggesting a possible redundancy of the system. However, an increased selectivity of the action of chemokines is thought to be gained through their ability to discern and preferentially bind to certain glycosaminoglycan (GAG) subpopulations, a process considered important in mediating tissue-specific leukocyte recruitment. [3][4][5][6] The chemokine RANTES has a complex influence on the biology of a variety of cell types including T lymphocytes, monocytes, natural killer cells, dendritic cells, basophils, and eosinophils. 7,8 At nanomolar concentrations, RANTES binds to and activates several 7-transmembrane G protein-coupled receptors (GPCRs), namely the chemokine receptors CCR1, CCR3, and CCR5. Ligation of these receptors activates a heterotrimeric G␣i protein-coupled signaling pathway, characterized by a transient Ca 2ϩ influx, 7,9 and triggers activation of cell polarization and chemotaxis. 10 In addition to these classic chemokine-activated responses, RANTES also induces several biochemical and biologic effects that are to date unique to this chemokine and that are triggered through a GPCR-independent pathway. Induction of this pathway is mediated by protein tyrosine kinases (PTKs), occurs at high micromolar concentrations of the chemokine, and leads to a sustained influx of Ca 2ϩ . 9 We and others have previously demonstr...

Recently, passive immunization of human immunodeficiency virus (HIV)-