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International audience; The 5′-untranslated region (5′-UTR) of the genomic RNA of human immunodeficiency viruses type-1 (HIV-1) and type-2 (HIV-2) is composed of highly structured RNA motifs essential for viral replication that are expected to interfere with Gag and Gag-Pol translation. Here, we have analyzed and compared the properties by which the viral 5′-UTR drives translation from the genomic RNA of both human immunodeficiency viruses. Our results showed that translation from the HIV-2 gRNA was very poor compared to that of HIV-1. This was rather due to the intrinsic structural motifs in their respective 5′-UTR without involvement of any viral protein. Further investigation pointed to a different role of TAR RNA, which was much inhibitory for HIV-2 translation. Altogether, these data highlight important structural and functional differences between these two human pathogens.

International audience; HIV-2 is less pathogenic for humans than HIV-1 and might provide partial cross-protection from HIV-1-induced pathology. Although both viruses replicate in the T cells of infected patients, only HIV-2 replicates efficiently in dendritic cells (DCs) and activates innate immune pathways. How HIV is sensed in DC is unknown. Capsid-mutated HIV-2 revealed that sensing by the host requires viral cDNA synthesis, but not nuclear entry or genome integration. The HIV-1 capsid prevented viral cDNA sensing up to integration, allowing the virus to escape innate recognition. In contrast, DCs sensed capsid-mutated HIV-1 and enhanced stimulation of T cells in the absence of productive infection. Finally, we found that DC sensing of HIV-1 and HIV-2 required the DNA sensor cGAS. Thus, the HIV capsid is a determinant of innate sensing of the viral cDNA by cGAS in dendritic cells. This pathway might potentially be harnessed to develop effective vaccines against HIV-1.

Type I interferons induce a complex transcriptional program that leads to a generalized antiviral response against a large panel of viruses, including human immunodeficiency virus type 1 (HIV-1). However, despite the fact that interferons negatively regulate HIV-1 ex vivo , a chronic interferon state is linked to the progression of AIDS and to robust viral replication, rather than protection, in vivo . To explain this apparent contradiction, we hypothesized that HIV-1 may have evolved a partial resistance to interferon, and to test this hypothesis, we analyzed the effects of alpha interferon (IFN-α) on the infectivity of HIV-1, human immunodeficiency virus type 2 (HIV-2), and rhesus monkey simian immunodeficiency virus (SIVmac). The results we obtained indicate that HIV-1 is more resistant to an IFN-α-induced response than are HIV-2 and SIVmac. Our data indicate that the accumulation of viral DNA is more compromised following the infection of IFN-α-treated cells with HIV-2 and SIVmac than with HIV-1. This defect correlates with a faster destabilization of HIV-2 viral nucleoprotein complexes (VNCs), suggesting a link between VNC destabilization and impaired viral DNA (vDNA) accumulation. The differential susceptibilities to IFN-α of the primate lentiviruses tested here do not map to the capsid protein (CA), excluding de facto a role for human tripartite motif protein isoform 5 alpha (Trim5α) in this restriction; this also suggests that an additional restriction mechanism differentially affects primate lentivirus infection. The different behaviors of HIV-1 and HIV-2 with respect to IFN-α responses may account at least in part for the differences in pathogenesis observed between these two virus types.

International audience; The human immunodeficiency virus (HIV) unspliced full length genomic RNA possesses features of an eukaryotic cellular mRNA as it is capped at its 5' end and polyadenylated at its 3' extremity. This genomic RNA is used both for the production of the viral structural and enzymatic proteins (Gag and Pol, respectively) and as genome for encapsidation in the newly formed viral particle. Although both of these processes are critical for viral replication, they should be controlled in a timely manner for a coherent progression into the viral cycle. Some of this regulation is exerted at the level of translational control and takes place on the viral 5' untranslated region and the beginning of the gag coding region. In this review, we have focused on the different initiation mechanisms (cap- and internal ribosome entry site (IRES)-dependent) that are used by the HIV-1 and HIV-2 genomic RNAs and the cellular and viral factors that can modulate their expression. Interestingly, although HIV-1 and HIV-2 share many similarities in the overall clinical syndrome they produce, in some aspects of their replication cycle, and in the structure of their respective genome, they exhibit some differences in the way that ribosomes are recruited on the gag mRNA to initiate translation and produce the viral proteins; this will be discussed in the light of the literature.

International audience; SAMHD1 is a newly identified restriction factor that targets lentiviruses in myeloid cells and is countered by the SIV(SM)/HIV-2 Vpx protein. By analyzing a large panel of Vpx mutants, we identify several residues throughout the 3-helix bundle predicted for Vpx that impair both its functionality and its ability to degrade SAMHD1. We determine that SAMHD1 is a strictly non-shuttling nuclear protein and that as expected WT Vpx localizes with it in the nucleus. However, we also identify a functional Vpx mutant with predominant cytoplasmic distribution that colocalizes with SAMHD1 in this location, suggesting that Vpx may also retain SAMHD1 in the cell cytoplasm, prior to its entry into the nucleus. Several mutations in Vpx were shown to affect the stability of Vpx, as well as Vpx:Vpx interactions. However, no strict correlation was observed between these parameters and the functionality of Vpx, implying that neither properties is absolutely required for this function and indicating that even unstable Vpx mutants may be very efficient in inducing SAMHD1 degradation. Overall, our analysis identifies several Vpx residues required for SAMHD1 degradation and points to a very efficient and plastic mechanism through which Vpx depletes this restriction factor.

International audience; OBJECTIVE: The authors had for aim to describe the effectiveness and the safety of a saquinavir/ritonavir (SQV/r) regimen, 1000/100mg twice daily, in HIV-infected pregnant patients. PATIENTS AND METHOD: We made a prospective and observational study of HIV positive female patients beginning or going on SQV/r antiretroviral treatment (ART) during pregnancy. RESULTS: Sixty-two patients were enrolled from July 2007 to June 2009 in 10 infectious diseases units in France. Thirty-six women (group 1) were ART naive on inclusion, 20 (group 2) had been previously treated and then switched to SQV/r, six (group 3) were treated with SQV/r before pregnancy. 58 patients delivered while on SQV/r regimen after a median pregnancy duration of 39 WA. Eighty percent had a viral load below 50 copies/mL and 93% below 400 copies/mL: respectively 77% and 93.5% in group 1, 83% and 89% in group 2, 83% and 100% in group 3. The median SQV minimum concentrations (C(min)) measured at the third trimester and at delivery were adequate, respectively 0.91 mg/L and 0.86 mg/L. Most women (52%) had a vaginal delivery; 12 (21%) had an elective caesarean section, for obstetrics factors in eight cases. None of the newborns were HIV-infected at 6 months of age (n = 59, one death at day 3). Only one severe adverse event occurred due to saquinavir (maternal grade 3 hepatotoxicity). CONCLUSION: SQV/r 1000/100mg twice daily seems to be effective and safe in HIV-infected pregnant women with adequate saquinavir C(min).

Objective To study the contribution of routine viral screening tests in patients with early rheumatoid arthritis (RA) or a potential for progressing to RA. Methods Eight hundred thirteen patients with swelling of at least 2 joints for at least 6 weeks and a symptom duration of less than 6 months in the ESPOIR cohort were screened for parvovirus B19 (IgG and IgM anti–parvovirus B19 antibodies), hepatitis B virus (HBV; hepatitis B surface antigen), hepatitis C virus (HCV; anti-HCV antibodies), and human immunodeficiency virus (HIV; anti–HIV-1 and -2 antibodies). Results Parvovirus B19 testing was performed in 806 patients and showed longstanding immunity in 574 (71.2%) and no antibodies in 223 (27.7%). Among the 9 remaining patients (7 IgG positive/IgM positive, 1 IgG negative/IgM positive, and 1 IgG indeterminate/IgM positive), only 2 (0.25%; 95% confidence interval [95% CI] 0–0.99%) had a positive polymerase chain reaction test for parvovirus B19; these patients (women ages 34 and 40 years) had no extraarticular signs. HIV seroprevalence was 0.12% (n = 1 of 813; 95% CI 0.01–0.8%) and HCV seroprevalence was 0.86% (n = 7 of 808, 95% CI 0.38–1.86%). HCV-related arthritis was diagnosed in 4 patients (0.5%). HCV-seropositive patients had significantly higher transaminase levels than the other patients (P = 0.001), with no significant differences for the other laboratory data. HBV seroprevalence was 0.12% (n = 1 of 808; 95% CI 0.01–0.8%); the positive HBV status was known before study inclusion, and the patient had no diagnosis of HBV-related arthritis. Finally, routine viral testing identified 2 patients with parvovirus B19 infection and 3 with HBV infection (0.6%; 95% CI 0.2–1.5%). Cost was €85.05 per patient (total €68,720). Conclusion Routine serologic testing did not contribute substantially to the diagnosis in this context.

Background HIV-1 is usually transmitted in the presence of semen. We have shown that semen boosts HIV-1 infection and contains fragments of prostatic acid phosphatase (PAP) forming amyloid aggregates termed SEVI (semen-derived enhancer of viral infection) that promote virion attachment to target cells. Despite its importance for the global spread of HIV-1, however, the effect of semen on virus infection is controversial. Results Here, we established methods allowing the meaningful analysis of semen by minimizing its cytotoxic effects and partly recapitulating the conditions encountered during sexual HIV-1 transmission. We show that semen rapidly and effectively enhances the infectivity of HIV-1, HIV-2, and SIV. This enhancement occurs independently of the viral genotype and coreceptor tropism as well as the virus producer and target cell type. Semen-mediated enhancement of HIV-1 infection was also observed under acidic pH conditions and in the presence of vaginal fluid. We further show that the potency of semen in boosting HIV-1 infection is donor dependent and correlates with the levels of SEVI. Conclusions Our results show that semen strongly enhances the infectivity of HIV-1 and other primate lentiviruses and that SEVI contributes to this effect. Thus, SEVI may play an important role in the sexual transmission of HIV-1 and addition of SEVI inhibitors to microbicides may improve their efficacy.

Background The 18 residue tail abutting the SH3 fold that comprises the heart of the C-terminal domain is the only part of HIV-1 integrase yet to be visualized by structural biology. To ascertain the role of the tail region in integrase function and HIV-1 replication, a set of deletion mutants that successively lacked three amino acids was constructed and analyzed in a variety of biochemical and virus infection assays. HIV-1/2 chimers, which harbored the analogous 23-mer HIV-2 tail in place of the HIV-1 sequence, were also studied. Because integrase mutations can affect steps in the replication cycle other than integration, defective mutant viruses were tested for integrase protein content and reverse transcription in addition to integration. The F185K core domain mutation, which increases integrase protein solubility, was furthermore analyzed in a subset of mutants. Results Purified proteins were assessed for in vitro levels of 3' processing and DNA strand transfer activities whereas HIV-1 infectivity was measured using luciferase reporter viruses. Deletions lacking up to 9 amino acids (1-285, 1-282, and 1-279) displayed near wild-type activities in vitro and during infection. Further deletion yielded two viruses, HIV-11-276 and HIV-11-273, that displayed approximately two and 5-fold infectivity defects, respectively, due to reduced integrase function. Deletion mutant HIV-11-270 and the HIV-1/2 chimera were non-infectious and displayed approximately 3 to 4-fold reverse transcription in addition to severe integration defects. Removal of four additional residues, which encompassed the C-terminal β strand of the SH3 fold, further compromised integrase incorporation into virions and reverse transcription. Conclusion HIV-11-270, HIV-11-266, and the HIV-1/2 chimera were typed as class II mutant viruses due to their pleiotropic replication defects. We speculate that residues 271-273 might play a role in mediating the known integrase-reverse transcriptase interaction, as their removal unveiled a reverse transcription defect. The F185K mutation reduced the in vitro activities of 1-279 and 1-276 integrases by about 25%. Mutant proteins 1-279/F185K and 1-276/F185K are therefore highlighted as potential structural biology candidates, whereas further deleted tail variants (1-273/F185K or 1-270/F185K) are less desirable due to marginal or undetectable levels of integrase function.

OBJECTIVES: To analyse the association between the presence of resistance mutations and treatment outcomes. The impact of HIV-1 drug resistance mutations in African adults on HAART has so far never been reported. METHODS: In 2004 in Abidjan, C?d'Ivoire, 106 adults on HAART had plasma viral load measurements. Patients with detectable viral loads had resistance genotypic tests. Patients were followed until 2006. Main outcomes were serious morbidity and immunological failure (CD4 cell count < 200 cells/microl). RESULTS: At study entry, the median previous time on HAART was 37 months and the median CD4 cell count was 266 cells/microl; 58% of patients had undetectable viral loads, 20% had detectable viral loads with no major resistance mutations, and 22% had detectable viral loads with one or more major mutations. The median change in CD4 cell count between study entry and study termination was +129 cells/microl in patients with undetectable viral loads, +51 cells/microl in those with detectable viral loads with no mutations and +3 cells/microl in those with detectable viral loads with resistance mutations. Compared with patients with undetectable viral loads, those with detectable viral loads with resistance mutations had adjusted hazard ratios of immunological failure of 4.32 (95%CI 1.38-13.57, P = 0.01). One patient died. The 18-month probability of remaining free of morbidity was 0.79 in patients with undetectable viral loads and 0.69 in those with resistance mutations (P = 0.19). CONCLUSION: In this setting with restricted access to second-line HAART, patients with major resistance mutations had higher rates of immunological failure, but most maintained stable CD4 cell counts and stayed alive for at least 20 months.

International audience; To report the tolerance of indinavir combined with ritonavir (IDV/r 800/100 mg) twice daily (bid) in sub-Saharan African HIV-infected adults. HAART-naives patients started zidovudine plus lamivudine plus IDV/r 800/100 mg bid. Follow-up included standardized documentation of morbidity, CD4(+) cell count, creatininemia, plasma HIV-1 RNA, and IDV minimal plasma concentration (C(min)) measurements at month 1 (M1), M3, and M6. Seventy HIV-1-infected adults (68 women, median CD4 235/mm(3)) started HAART. At M6, 63% had undetectable viral load, and the median gain in CD4 since baseline was +128/mm(3). During the first 6 months, 21 patients experimented with 23 treatment modifications (reduction in IDV/r 400/100 mg bid, n = 11; switch to efavirenz, n = 11; zidovudine replaced by stavudine, n = 1), including 22 for digestive intolerance and 1 for severe anemia. At M1, M3, and M6, 67, 59, and 48 patients were still receiving IDV/r 800/100 mg bid, of whom 70%, 72%, and 60% had IDV Cmin above 5 ng/ml, respectively. In these patients, at M1, M3, and M6, the mean (+/- SD) IDV C(min) were 3431 +/- 3835 ng/ml, 2288 +/- 2116 ng/ml, and 1543 +/- 2398 ng/ml, respectively. There was no renal insufficiency of any grade, and no symptoms of urinary stones. The IDV/r 800/100 mg bid-containing regimen led to high IDV Cmin and a high rate of digestive intolerance. There was a surprising lack of nephrological side effects during the 6 months of follow-up, supporting the hypothesis that nephrological tolerance of IDV might be higher in sub-Saharan African individuals than in Americans or Europeans.

International audience; The dimerization processes of the human immunodeficiency virus (HIV) types 1 and 2 reverse transcriptase (RTs) from their subunits have been investigated using a number of complementary approaches (fluorescence spectroscopy, size exclusion-HPLC and polymerase activity assay). The formation of the native heterodimeric form of HIV-1 and HIV-2 RT occurs in a two step process. The first step is a concentration-dependent association of the two subunits (p66 and p51) to give a heterodimeric intermediate, which slowly isomerizes to the "mature" heterodimeric form of the enzyme. For both RTs, the first step behaves as a second order reaction with similar association rate constants (in the range of 2 x 10(4) to 4 x 10(4) M-1 s-1). This initial dimerization results in a 25% quenching of the intrinsic fluorescence and a 30% decrease in the accessibility of the tryptophan hydrophobic cluster to solvent as revealed by iodide quenching experiments and by monitoring the binding of 1-anilino-8-naphthalenesulphonate. The formation of the intermediate-RT form appears to involve hydrophobic regions of the subunits containing tryptophan residues. This intermediate form is devoid of polymerase activity, but is able to bind primer/template with high affinity. The final stage of the mature RT-heterodimer formation occurs in a slow first order reaction, which is 12-fold faster for HIV-2 (1.2 h-1) than HIV-1 RT (0.1 h-1). At micromolar concentrations, this slow isomerization constitutes the rate limiting step of the RT maturation and the structural change involved appears to be partly associated with the catalytic site, as shown using fluorescent labelled primer/template. On the basis of both the presently available X-ray structure of the HIV-1 RT and the predicted structure of HIV-2 RT, the thumb subdomain of the p51 subunit seems to be involved in this maturation step, which is probably the interaction of this domain with the RNAse H domain of the large subunit. The placement of the fingers subdomain of p51 in the palm subdomain of the p66 subunit may also be associated with formation of mature heterodimeric RTs.