High mutation frequency during reverse transcription has a principal role in the genetic variation of primate lentiviral populations. It is the main driving force for the generation of drug resistance and the escape from immune surveillance. G to A hypermutation is one of the characteristics of primate lentiviruses, as well as other retroviruses, during replication in vivo and in cell culture 1-6 . The molecular mechanisms of this process, however, remain to be clarified. Here, we demonstrate that CEM15 (also known as apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G; APOBEC3G) 7,8 , an endogenous inhibitor of human immunodeficiency virus type 1 (HIV-1) replication, is a cytidine deaminase and is able to induce G to A hypermutation in newly synthesized viral DNA. This effect can be counteracted by the HIV-1 virion infectivity factor (Vif). It seems that this viral DNA mutator is a viral defence mechanism in host cells that may induce either lethal hypermutation or instability of the incoming nascent viral reverse transcripts, which could account for the Vif-defective phenotype. Importantly, the accumulation of CEM15-mediated non-lethal hypermutation in the replicating viral genome could potently contribute to the genetic variation of primate lentiviral populations.HIV-1 Vif protein is required for viral replication in vivo and in some 'non-permissive' cells, such as peripheral blood mononuclear cells, macrophages and H9 T cells 9-11 . The vifdefective viruses (Δvif) from non-permissive cells cannot complete reverse transcription, or the newly synthesized DNA cannot exist in the target cells for a significant time period 12-14 . Recently, it has been demonstrated that CEM15 is an endogenous inhibitor of HIV-1 that exists only in non-permissive cells. Its inhibitory effect on HIV-1 replication can be counteracted by HIV-1 Vif protein 7 . As Vif binds to HIV-1 RNA in the cytoplasm of virusproducing cells 15-17 , we investigated whether CEM15, which shares significant homology with some other cytidine deaminases that edit RNA, could also edit HIV-1 genomic or spliced RNA. We have sequenced the nearly full-length genomic RNA (>98%) of HIV-1 from the Δvif virions generated from H9 T cells by polymerase chain reaction with reverse transcription (RT-PCR) techniques (primer pairs are listed in Supplementary Table S1). Compared with the sequence of pNL4-3Δvif DNA, the change of genomic RNA in the virions is not significant. We have found an A to G change at positions 2257 and 3608, and a G to T change at position 9418 (data not shown). We have also sequenced several spliced HIV-1(NL4-3Δvif) RNA in H9 cells ( Supplementary Fig. S1 and Table S1); however, no mutations were demonstrated.Correspondence and requests for materials should be addressed to H.Z (hui.zhang@jefferson.edu).. Supplementary Information accompanies the paper on www.nature.com/nature. Competing interests statementThe authors declare that they have no competing financial interests. Table S1) that we have analysed (Fig. 1a, row 2 ...

The interferon (IFN) system, including various IFNs and IFN-inducible gene products, is well known for its potent innate immunity against wide-range viruses. Recently, a family of cytidine deaminases, functioning as another innate immunity against retroviral infection, has been identified. However, its regulation remains largely unknown. In this report, we demonstrate that through a regular IFN-␣/␤ signal transduction pathway, IFN-␣ can significantly enhance the expression of apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (APOBEC3G) in human primary resting but not activated CD4 T cells and the amounts of APOBEC3G associated with a low molecular mass. Interestingly, short-time treatments of newly infected resting CD4 T cells with IFN-␣ will significantly inactivate human immunodeficiency virus type 1 (HIV-1) at its early stage. This inhibition can be counteracted by APOBEC3G-specific short interfering RNA, indicating that IFN-␣-induced APOBEC3G plays a key role in mediating this anti-HIV-1 process. Our data suggest that APOBEC3G is also a member of the IFN system, at least in resting CD4 T cells. Given that the IFN-␣/ APOBEC3G pathway has potent anti-HIV-1 capability in resting CD4 T cells, augmentation of this innate immunity barrier could prevent residual HIV-1 replication in its native reservoir in the post-highly active antiretroviral therapy era.

No abstract

Antimicrobial peptides (AMPs) have gained increasing attention, as they can overcome recurring microbial invasions. However, their poor antimicrobial activity and potential cytotoxicity remain impediments to their clinical applications as novel therapeutic agents. To enhance the antimicrobial activity and cell selectivity of AMPs, a series of amphiphilic peptides based on leucocin A were designed by substituting noncharged hydrophilic residues with arginine and leucine. Of the engineered peptides, peptide 7 (WRL3) (WLRAFRRLVRRLARGLRR-NH2) exhibited the highest cell selectivity toward bacterial cells over erythrocytes and macrophages. Fluorescent measurements and microscopic observations demonstrated that 7 increased cell membrane permeability and disrupted membrane envelope integrity, and eventually led to whole cell lysis. Additionally, flow cytometry analysis and subcellular localization studies revealed that 7 showed potent cytotoxicity against human hepatoma cells HepG2. In summary, the data indicate that these engineered peptides, in particular 7, have enormous promise for antibacterial and/or antitumor therapeutics.

BackgroundBacillus subtilis strain PB2-L1 produces the lipopeptide surfactin, a highly potent biosurfactant synthesized by a large multimodular nonribosomal peptide synthetase (NRPS). In the present study, the modules SrfA-A-Leu, SrfA-B-Asp, and SrfA-B-Leu from surfactin NRPS in B. subtilis BP2-L1 were successfully knocked-out using a temperature-sensitive plasmid, pKS2-mediated-based, homologous, recombination method.ResultsThree novel surfactin products were produced, individually lacking amino acid Leu-3, Asp-5, or Leu-6. These surfactins were detected, isolated, and characterized by HPLC and LC-FTICR-MS/MS. In comparison with native surfactin, [∆Leu3]surfactin and [∆Leu6]surfactin showed evidence of reduced toxicity, while [∆Asp5]surfactin showed stronger inhibition than native surfactin against B. pumilus and Micrococcus luteus. These results showed that the minimum inhibitory concentration of [∆Leu6]surfactin for Fusarium moniliforme was 50 μg/mL, such that [∆Leu6]surfactin could lead to mycelium projection, cell damage, and leakage of nucleic acids and protein. These factors all contributed to stimulating apoptosis in F. moniliforme.ConclusionsThe present results revealed that [∆Leu6]surfactin showed a significant antifungal activity against F. moniliforme and might successfully be employed to control fungal food contamination and improve food safety.

Methicillin-resistant Staphylococcus aureus (MRSA) has become increasingly prevalent in hospitals, clinics, and the community. MRSA can cause significant and even lethal infections, especially in skin burn wounds. The currently available topical agents have largely failed to eliminate MRSA infections due to resistance. Therefore, there is an urgent need for new and effective approaches for treating MRSA. Here, we show that a novel engineered amphipathic peptide, WRL3 (WLRAFRRLVRRLARGLRR-NH2), exhibits potent antimicrobial activity against MRSA, even in the presence of various salts or serum. The cell selectivity of WRL3 was demonstrated by its ability to specifically eliminate MRSA cells over host cells in a coculture model. Additionally, WRL3 showed a synergistic effect against MRSA when combined with ceftriaxone and effectively inhibited sessile biofilm bacteria growth leading to a reduction in biomass. Fluorescent measurements and microscopic observations of live bacterial cells and artificial membranes revealed that WRL3 exerted its bactericidal activity possibly by destroying the bacterial membrane. In vivo studies indicate that WRL3 is able to control proliferation of MRSA in wound tissue and reduce bioburden and provides a more favorable environment for wound healing. Collectively, our data suggest that WRL3 has enormous potential as a novel antimicrobial agent for the treatment of clinical MRSA infections of skin burn wounds.

To convert the lipopeptide non-producer strain Bacillus subtilis pB2 into a plipastatin and surfactin coproducer, a gene expression cassette composed of a constitutive promoter (P43), functional gene sfp, and pleiotropic regulatory gene degQ was integrated into the chromosomal amyE locus of strain B. subtilis pB2 by homologous recombination, which generated a plipastatin and surfactin co-producer. Thirteen plipastatins and fifteen surfactins were identified in lipopeptide extracts using analytical techniques, and their effects on microorganisms were described by microscopic, cytoskeleton analysis and flow-cytometry, respectively. Plipastatins isolated from the engineered strain pB2-L exhibited strong antifungal activity (MIC 16 μg ml) by disrupting the cell walls, membranes and cytoskeleton of Fusarium oxysporum f. sp. cucumerinum hyphae. Surfactins affected the cell membrane of Staphylococcus aureus (MIC 20 μg ml), resulting in nucleic acid leakage and ultimately, cell death. Based on the convenience of genetic manipulation in the engineering strain, this work could be useful for the rational design of lipopeptide synthetases via the recombination of gene fragments to generate arrays of peptide derivatives and thus expand the diversity of microbial-produced lipopeptides.

Dopa and lysine are widely found in mussel foot proteins and are suggested to play synergistic roles in wet adhesion; yet, the detailed molecular mechanism remains unclear.