Identification of a novel phosphorylation site in hepatitis C virus NS5A

Hepatitis C virus (HCV) NS5A protein is phosphorylated on multiple residues; however, despite extensive study, the precise identity of these sites has not been determined unambiguously. In this study, we have used a combination of immunoprecipitation and mass spectrometry to identify these phosphorylation sites. This analysis revealed the presence of a major phosphorylated residue within NS5A from the genotype 1b Con1 isolate – serine 249 (serine 2221 in polyprotein numbering). However, mutation of this residue (or the corresponding threonine in the JFH-1 isolate) to either a phosphomimetic (aspartate) or a phosphoablative (alanine) residue resulted in no phenotype. We conclude that phosphorylation of this residue, in the context of a highly culture-adapted HCV genome, does not play a role in either viral RNA replication or virus assembly. It is possible that it might be important in an aspect of virus biology that is not recapitulated faithfully in the Huh-7 cell-culture system

<i>TP53</i> drives invasion through expression of its ∆133p53β variant

International audienceTP53 is conventionally thought to prevent cancer formation and progression to metastasis, while mutant TP53 has transforming activities. However, in the clinic, TP53 mutation status does not accurately predict cancer progression. Here we report, based on clinical analysis corroborated with experimental data, that the p53 isoform D133p53b promotes cancer cell invasion, regardless of TP53 mutation status. D133p53b increases risk of cancer recurrence and death in breast cancer patients. Furthermore D133p53b is critical to define invasiveness in a panel of breast and colon cell lines, expressing WT or mutant TP53. Endogenous mutant D133p53b depletion prevents invasiveness without affecting mutant full-length p53 protein expression. Mechanistically WT and mutant D133p53b induces EMT. Our findings provide explanations to 2 long-lasting and important clinical conundrums: how WT TP53 can promote cancer cell invasion and reciprocally why mutant TP53 gene does not systematically induce cancer progression

The subcellular localization of the hepatitis C virus non-structural protein NS2 is regulated by an ion channel-independent function of the p7 protein

The hepatitis C virus (HCV) p7 ion channel and non-structural protein 2 (NS2) are both required for efficient assembly and release of nascent virions, yet precisely how these proteins are able to influence this process is unclear. Here, we provide both biochemical and cell biological evidence for a functional interaction between p7 and NS2. We demonstrate that in the context of a genotype 1b subgenomic replicon the localization of NS2 is affected by the presence of an upstream p7 with its cognate signal peptide derived from the C terminus of E2 (SPp7). Immunofluorescence analysis revealed that the presence of SPp7 resulted in the targeting of NS2 to sites closely associated with viral replication complexes. In addition, biochemical analysis demonstrated that, in the presence of SPp7, a significant proportion of NS2 was found in a detergent (Triton X-100)-insoluble fraction, which also contained a marker of detergent resistant rafts. In contrast, in replicons lacking p7, NS2 was entirely detergent soluble and the altered localization was lost. Furthermore, we found that serine 168 within NS2 was required for its localization adjacent to replication complexes, but not for its accumulation in the detergent-insoluble fraction. NS2 physically interacted with NS5A and this interaction was dependent on both p7 and serine 168 within NS2. Mutational and pharmacological analyses demonstrated that these effects were not a consequence of p7 ion channel function, suggesting that p7 possesses an alternative function that may influence the coordination of virus genome replication and particle assembly

Inhibition of human gastric and pancreatic lipases by chiral alkylphosphonates. A kinetic study with 1,2-didecanoyl-sn-glycerol monolayer

International audienc

The V domain of herpesvirus Ig-like receptor (HIgR) contains a major functional region in herpes simplex virus-1 entry into cells and interacts physically with the viral glycoprotein D

The herpesvirus entry mediator C (HveC), previously known as poliovirus receptor-related protein 1 (PRR1), and the herpesvirus Ig-like receptor (HIgR) are the bona fide receptors employed by herpes simplex virus-1 and -2 (HSV-1 and -2) for entry into the human cell lines most frequently used in HSV studies. They share an identical ectodomain made of one V and two C2 domains and differ in transmembrane and cytoplasmic regions. Expression of their mRNA in the human nervous system suggests possible usage of these receptors in humans in the path of neuron infection by HSV. Glycoprotein D (gD) is the virion component that mediates HSV-1 entry into cells by interaction with cellular receptors. We report on the identification of the V domain of HIgR/PRR1 as a major functional region in HSV-1 entry by several approaches. First, the epitope recognized by mAb R1.302 to HIgR/PRR1, capable of inhibiting infection, was mapped to the V domain. Second, a soluble form of HIgR/PRR1 consisting of the single V domain competed with cell-bound full- length receptor and blocked virion infectivity. Third, the V domain was sufficient to mediate HSV entry, as an engineered form of PRR1 in which the two C2 domains were deleted and the V domain was retained and fused to its transmembrane and cytoplasmic regions was still able to confer susceptibility, although at reduced efficiency relative to full-length receptor. Consistently, transfer of the V domain of HIgR/PRR1 to a functionally inactive structural homologue generated a chimeric receptor with virus-entry activity. Finally, the single V domain was sufficient for in vitro physical interaction with gD. The in vitro binding was specific as it was competed both by antibodies to the receptor and by a mAb to gD with potent neutralizing activity for HSV-1 infectivity