• Epigenetics reader
• Oncogenesis
• PSIP1
• Targeted therapy

• Humans
• Carcinogenesis/genetics
• Neoplasms/genetics
• Neoplasms/pathology
• Neoplasms/metabolism
• Neoplasms/drug therapy
• Adaptor Proteins, Signal Transducing/genetics
• Adaptor Proteins, Signal Transducing/metabolism
• Animals
• Gene Expression Regulation, Neoplastic
• Intercellular Signaling Peptides and Proteins/genetics
• Intercellular Signaling Peptides and Proteins/metabolism
• Transcription Factors/genetics
• Transcription Factors/metabolism

• ALLINI
• HIV-1 integrase
• antiretroviral drug
• pirmitegravir

• HIV-1/drug effects
• HIV-1/genetics
• HIV-1/enzymology
• Drug Resistance, Viral/genetics
• HIV Integrase/metabolism
• HIV Integrase/genetics
• HIV Integrase/chemistry
• HIV Integrase Inhibitors/pharmacology
• HIV Integrase Inhibitors/chemistry
• Humans
• HIV Infections/virology
• HIV Infections/drug therapy
• Mutation

• S10 OD028725 NIH HHS
• P30 AI050409 NIAID NIH HHS
• R01 AI141327 NIAID NIH HHS
• R01 AI184419 NIAID NIH HHS
• P20 GM103446 NIGMS NIH HHS
• CC2058 Arthritis Research UK
• Wellcome Trust
• AI 141327 HHS | NIH | National Institute of Allergy and Infectious Diseases (NIAID)
• R01 MH116695 NIMH NIH HHS
• U54 AI170791 NIAID NIH HHS
• U54 AI170855 NIAID NIH HHS

• Antiviral Agents/pharmacology
• Antiviral Agents/chemistry
• Drug Design
• Protein Multimerization/drug effects
• Humans
• Viral Proteins/chemistry
• Viral Proteins/antagonists & inhibitors
• Viral Proteins/metabolism
• SARS-CoV-2/drug effects
• Quality Control
• COVID-19 Drug Treatment

• ALLINI
• HIV
• aberrant integrase multimerization
• allosteric integrase inhibitor
• integrase
• quinoline
• virus maturation

• HIV-1/metabolism
• Allosteric Regulation
• HIV Integrase Inhibitors/pharmacology
• HIV Integrase Inhibitors/chemistry
• HIV Integrase/metabolism
• Quinolines/pharmacology
• Protein Multimerization

• P20 GM103476 NIGMS NIH HHS
• R01 AI140985 NIAID NIH HHS
• AI140985 NIH HHS

• ALLINI
• HIV-1 integrase
• Pirmitegravir
• antiretroviral drug

• S10 OD028725 NIH HHS
• P30 AI050409 NIAID NIH HHS
• R01 AI143649 NIAID NIH HHS
• R01 AI141327 NIAID NIH HHS
• P20 GM103446 NIGMS NIH HHS
• CC2058 Arthritis Research UK
• Wellcome Trust
• R01 MH116695 NIMH NIH HHS
• U54 AI170791 NIAID NIH HHS
• U54 AI170855 NIAID NIH HHS

• intrinsically disordered proteins
• molecular scaffolds
• structural biology
• transcription

• retrovirus
• innate immunity
• restriction factors

• H3K36me3
• HIV
• PIC
• PTM
• chromatin
• histone
• human immunodeficiency virus
• intasome
• integration
• nucleosome
• posttranslational modification
• preintegration complex

In infectious HIV-1 particles, the capsid protein (CA) forms a cone-shaped shell called the capsid, which encases the viral ribonucleoprotein complex (vRNP). Following cellular entry, the capsid is disassembled through a poorly understood process referred to as uncoating, which is required to release the reverse transcribed HIV-1 genome for integration into host chromatin. Whereas single virus imaging using indirect CA labeling techniques suggested uncoating to occur in the cytoplasm or at the nuclear pore, a recent study using eGFP-tagged CA reported uncoating in the nucleus. To delineate the HIV-1 uncoating site, we investigated the mechanism of eGFP-tagged CA incorporation into capsids and the utility of this fluorescent marker for visualizing HIV-1 uncoating. We find that virion incorporated eGFP-tagged CA is effectively excluded from the capsid shell, and that a subset of the tagged CA is vRNP associated. These results thus imply that eGFP-tagged CA is not a direct marker for capsid uncoating. We further show that native CA co-immunoprecipitates with vRNP components, providing a basis for retention of eGFP-tagged and untagged CA by sub-viral complexes in the nucleus. Moreover, we find that functional viral replication complexes become accessible to integrase-interacting host factors at the nuclear pore, leading to inhibition of infection and demonstrating capsid permeabilization prior to nuclear import. Finally, we find that HIV-1 cores containing a mixture of wild-type and mutant CA interact differently with cytoplasmic versus nuclear pools of the CA-binding host cofactor CPSF6. Our results suggest that capsid remodeling (including a loss of capsid integrity) is the predominant pathway for HIV-1 nuclear entry and provide new insights into the mechanism of CA retention in the nucleus via interaction with vRNP components.

The timing, location and mechanisms of HIV-1 capsid disassembly which is referred to as uncoating remains unclear. Direct labeling of HIV-1 capsids, by incorporating a few green fluorescent proteins (GFP) tagged capsid protein (CA) into virions allows to image the spatio-temporal loss of HIV-1 CA during virus infection. However, the localization and functions of a few virion incorporated eGFP-tagged CA proteins remain unclear, since <50% of virus packaged CA proteins participate to form the conical capsid shell that protects the HIV-1 genome. Here we developed several approaches to test the localization and function of eGFP-tagged CA proteins in virions. We found that eGFP-tagged CA proteins are excluded from the conical capsid shell and that a subset of these proteins is associated with the viral ribonucleoprotein complex (vRNPs), through direct interactions between CA and vRNP components. eGFP-tagged CA is retained in the nucleus by virtue of vRNP association and is unlikely to report on HIV-1 capsid disassembly. We also found that HIV-1 capsids become permeabilized and are remodeled during their transport into the nucleus. Our study provides new insights into the ability of CA to interact with vRNPs for its retention in the nucleus and highlights capsid remodeling as a preferred pathway for HIV-1 entry into the nucleus.

Allosteric HIV-1 integrase (IN) inhibitors, or ALLINIs, are a new class of antiviral agents that bind at the dimer interface of the IN, away from the enzymatic catalytic site and block viral replication by triggering an aberrant multimerization of the viral enzyme. To further our understanding of the important binding features of multi-substituted quinoline-based ALLINIs, we have examined the IN multimerization and antiviral properties of substitution patterns at the 6 or 8 position. We found that the binding properties of these ALLINIs are negatively impacted by the presence of bulky substitutions at these positions. In addition, we have observed that the addition of bromine at either the 6 (6-bromo) or 8 (8-bromo) position conferred better antiviral properties. Finally, we found a significant loss of potency with the 6-bromo when tested with the ALLINI-resistant IN A128T mutant virus, while the 8-bromo analog retained full effectiveness.

• HIV
• aberrant integrase multimerization
• allosteric integrase inhibitor
• integrase
• integrase-RNA binding
• virus maturation
• virus morphogenesis

• HIV Integrase/metabolism
• HIV Integrase Inhibitors/pharmacology
• HIV-1/genetics
• HIV-1/metabolism
• Humans
• RNA, Viral/genetics
• RNA, Viral/metabolism

• R01 AI070042 NIAID NIH HHS
• R01 AI143649 NIAID NIH HHS
• U54 AI150472 NIAID NIH HHS
• U54AI150472; R01AI070042; R01AI143649 NIAID NIH HHS

• Antiretroviral therapy
• Drug design
• Enzyme
• Hydrolase
• Retrovirus

• ledgf
• psip1
• hrp2
• hdgfl2
• hdgfrp2
• pro-seq
• pausing
• release

• Adaptor Proteins, Signal Transducing/chemistry
• Adaptor Proteins, Signal Transducing/metabolism
• Cell Line, Tumor
• DNA-Binding Proteins/chemistry
• DNA-Binding Proteins/metabolism
• Gene Expression
• Humans
• Intrinsically Disordered Proteins/chemistry
• Intrinsically Disordered Proteins/metabolism
• Models, Molecular
• Mutation
• Protein Binding
• Protein Domains
• Protein Interaction Domains and Motifs/genetics
• Protein Interaction Maps
• RNA Polymerase II/chemistry
• RNA Polymerase II/metabolism
• RNA-Binding Proteins/chemistry
• RNA-Binding Proteins/genetics
• RNA-Binding Proteins/metabolism
• Transcription Elongation, Genetic
• Transcription Factors/chemistry
• Transcription Factors/genetics
• Transcription Factors/metabolism
• Transcriptional Elongation Factors/chemistry
• Transcriptional Elongation Factors/metabolism

• R35 GM137996 NIGMS NIH HHS
• T32 ES027801 NIEHS NIH HHS

• 5′-UTR
• ALLINI
• HIV-1
• Integrase
• Nucleocapsid
• RNA binding
• RNA secondary structure
• XL-SHAPE

• Clonal dynamics
• Gene therapy
• HSPC
• Humanized mouse
• LoVIS-Seq
• MDA
• VIS tracking

• Animals
• Disease Models, Animal
• Hematopoiesis
• Hematopoietic Stem Cell Transplantation
• Hematopoietic Stem Cells
• Humans
• Mice
• Mice, Inbred NOD
• Mice, SCID

• AI110297 NIH HHS
• R00 HL116234 NHLBI NIH HHS
• HL126544 NIH HHS
• HL125030 NIH HHS
• R01 AI145038 NIAID NIH HHS
• R56 HL126544 NHLBI NIH HHS
• R01 HG010318 NHGRI NIH HHS
• R21 HG010108 NHGRI NIH HHS
• AI145038 NIH HHS
• K99 HL116234 NHLBI NIH HHS
• 5U19AI117941 NIH HHS
• national institutes of health
• california institute for regenerative medicine

• Awards and Prizes
• HIV Infections/virology
• HIV-1/genetics
• HIV-1/physiology
• History, 20th Century
• History, 21st Century
• Human Immunodeficiency Virus Proteins/genetics
• Human Immunodeficiency Virus Proteins/metabolism
• Humans
• Virology/history

• hiv
• ledgin
• antiviral agents
• block and lock
• functional cure
• integration

During the integration step, human immunodeficiency virus type 1 integrase (IN) interacts with viral DNA and the cellular cofactor LEDGF/p75 to effectively integrate the reverse transcript into the host chromatin. Allosteric human immunodeficiency virus type 1 integrase inhibitors (ALLINIs) are a new class of antiviral agents that bind at the dimer interface of the IN catalytic core domain and occupy the binding site of LEDGF/p75. While originally designed to block IN-LEDGF/p75 interactions during viral integration, several of these compounds have been shown to also severely impact viral maturation through an IN multimerization mechanism. In this study, we tested the hypothesis that these dual properties of ALLINIs could be decoupled toward late stage viral replication effects by generating additional contact points between the bound ALLINI and a third subunit of IN. By sequential derivatization at position 7 of a quinoline-based ALLINI scaffold, we show that IN multimerization properties are enhanced by optimizing hydrophobic interactions between the compound and the C-terminal domain of the third IN subunit. These features not only improve the overall antiviral potencies of these compounds but also significantly shift the ALLINIs selectivity toward the viral maturation stage. Thus, we demonstrate that to fully maximize the potency of ALLINIs, the interactions between the inhibitor and all three IN subunits need to be simultaneously optimized.

• ALLINI
• antiviral agent
• drug design
• human immunodeficiency virus
• integrase
• oligomerization
• protein–protein interaction
• quinoline

• AIDS/HIV
• Drug resistance
• Entry inhibitors
• Integrase inhibitor
• Non-nucleoside RT inhibitors
• Nucleoside RT inhibitors
• Protease inhibitors
• X-ray crystallography

• CPSF6
• LEDGF/p75
• human immunodeficiency virus
• integration
• lentiviruses

• Animals
• Cats/genetics
• Cats/virology
• Cattle/genetics
• Cattle/virology
• Cell Line
• Evolution, Molecular
• HEK293 Cells
• Horses/genetics
• Horses/virology
• Humans
• Intercellular Signaling Peptides and Proteins/genetics
• Jurkat Cells
• Lentivirus/genetics
• Macaca mulatta/genetics
• Macaca mulatta/virology
• Mice/genetics
• Mice/virology
• Primates/genetics
• Primates/virology
• Virus Integration/genetics
• Virus Replication
• mRNA Cleavage and Polyadenylation Factors/genetics

• T32 AI007386 NIAID NIH HHS
• R37 AI039394 NIAID NIH HHS
• DP1 DA043915 NIDA NIH HHS
• R01 AI039394 NIAID NIH HHS
• R01 AI052014 NIAID NIH HHS
• P30 AI060354 NIAID NIH HHS
• HHS | NIH | National Institute of Allergy and Infectious Diseases (NIAID)

HIV-1 persists lifelong in memory cells of the immune system as latent provirus that rebounds upon treatment interruption. Therefore, the latent reservoir is the main target for an HIV cure. Here, we studied the direct link between integration site and transcription using LEDGINs and Barcoded HIV-ensembles (B-HIVE). LEDGINs are antivirals that inhibit the interaction between HIV-1 integrase and the chromatin-tethering factor LEDGF/p75. They were used as a tool to retarget integration, while the effect on HIV expression was measured with B-HIVE. B-HIVE tracks insert-specific HIV expression by tagging a unique barcode in the HIV genome. We confirmed that LEDGINs retarget integration out of gene-dense and actively transcribed regions. The distance to H3K36me3, the marker recognized by LEDGF/p75, clearly increased. LEDGIN treatment reduced viral RNA expression and increased the proportion of silent provirus. Finally, silent proviruses obtained after LEDGIN treatment were located further away from epigenetic marks associated with active transcription. Interestingly, proximity to enhancers stimulated transcription irrespective of LEDGIN treatment, while the distance to H3K36me3 only changed after treatment with LEDGINs. The fact that proximity to these markers are associated with RNA expression support the direct link between provirus integration site and viral expression.

• intasome
• integrase
• integrase strand transfer inhibitor (INSTI)
• integration
• nucleoprotein complex
• retrovirus
• site-specific recombination

• AIDS
• HIV/AIDS
• antiretroviral therapy
• intasome
• integrase
• integrase strand transfer inhibitor
• integration
• microbiology
• polynucleotidyl transferase
• retrovirus
• viral DNA
• virology
• virus structure

The Moloney murine leukemia virus (MLV) is a prototype gammaretrovirus requiring nuclear disassembly before DNA integration. In the nucleus, integration site selection towards promoter/enhancer elements is mediated by the host factor bromo- and extraterminal domain (BET) proteins (bromodomain (Brd) proteins 2, 3 and 4). MLV-based retroviral vectors are used in gene therapy trials. In some trials leukemia occurred through integration of the MLV vector in close proximity to cellular oncogenes. BET-mediated integration is poorly understood and the nature of integrase oligomers heavily debated. Here, we created wild-type infectious MLV vectors natively incorporating fluorescent labeled IN and performed single-molecule intensity and Förster resonance energy transfer experiments. The nuclear localization of the MLV pre-integration complex neither altered the IN content, nor its quaternary structure. Instead, BET-mediated interaction of the MLV intasome with chromatin in the post-mitotic nucleus reshaped its quaternary structure.

To fulfill a productive infection cycle the human immunodeficiency virus (HIV) relies on host-cell factors. Interference with these co-factors holds great promise in protecting cells against HIV infection. LEDGF/p75, encoded by the PSIP1 gene, is used by the integrase (IN) protein in the pre-integration complex of HIV to bind host-cell chromatin facilitating proviral integration. LEDGF/p75 depletion results in defective HIV replication. However, as part of its cellular function LEDGF/p75 tethers cellular proteins to the host-cell genome. We used site-specific editing of the PSIP1 locus using CRISPR/Cas to target the aspartic acid residue in position 366 and mutated it to asparagine (D366N) to disrupt the interaction with HIV IN but retain LEDGF/p75 cellular function. The resulting cell lines demonstrated successful disruption of the LEDGF/p75 HIV-IN interface without affecting interaction with cellular binding partners. In line with LEDGF/p75 depleted cells, D366N cells did not support HIV replication, in part due to decreased integration efficiency. In addition, we confirm the remaining integrated provirus is more silent. Taken together, these results support the potential of site-directed CRISPR/Cas9 mediated knock-in to render cells more resistant to HIV infection and provides an additional strategy to protect patient-derived T-cells against HIV-1 infection as part of cell-based therapy.

• LEDGF/p75
• disordered proteins
• leukemia
• phosphorylation
• protein–protein interactions

• Adaptor Proteins, Signal Transducing/genetics
• Adaptor Proteins, Signal Transducing/metabolism
• Amino Acid Motifs
• Cell Line, Tumor
• HIV/enzymology
• HIV/genetics
• HIV Integrase/genetics
• HIV Integrase/metabolism
• Histone-Lysine N-Methyltransferase/genetics
• Histone-Lysine N-Methyltransferase/metabolism
• Humans
• Mediator Complex Subunit 1/genetics
• Mediator Complex Subunit 1/metabolism
• Myeloid-Lymphoid Leukemia Protein/genetics
• Myeloid-Lymphoid Leukemia Protein/metabolism
• Phosphorylation/genetics
• Transcription Factors/genetics
• Transcription Factors/metabolism

• R00 CA187565 NCI NIH HHS

• CPSF6
• Capsid
• HIV/AIDS
• Integrase
• LEDGF/p75
• Latency
• Viral DNA integration
• Virus–host interaction

• Gene Targeting
• HIV Integrase/physiology
• HIV-1/genetics
• HIV-1/physiology
• Host-Pathogen Interactions/genetics
• Humans
• Virus Integration/physiology
• Virus Internalization
• Virus Latency/physiology

• AI052014 National Institute of Allergy and Infectious Diseases
• R37 AI039394 NIAID NIH HHS
• R01 AI039394 NIAID NIH HHS
• R01 AI052014 NIAID NIH HHS
• AI039394 National Institute of Allergy and Infectious Diseases

The integrase strand-transfer inhibitors (INSTIs) are an important class in the arsenal of antiretroviral drugs designed to block the integration of HIV-1 cDNA into the host DNA through the inhibition of DNA strand transfer. In this study for the first time in Morocco, the complete HIV-1 integrase gene was analysed from newly diagnosed patients to evaluate the prevalence of natural polymorphisms and INSTIs resistance-associated mutations in the integrase gene.

The 864pb IN coding region was successfully sequenced from plasma sample for 77 among 80 antiretroviral naïve patients. The sequences were interpreted for drug resistance according to the Stanford algorithm. Sixty samples were HIV-1 subtype B (78%), fourteen CRF02_AG (18%), two subtype C and one subtype A. Overall 81 of 288 (28%) amino acid IN positions presented at least one polymorphism each. We found 18 (36.73%), 42 (25.76%) and 21 (27.27%) of polymorphic residues assigned to the N-Terminal Domain, Catalytic Core Domaine and the C-Terminal Domain positions respectively. Primary INSTIs resistance mutation were absent, however secondary mutations L74IM, T97A were detected in four samples (5.2%). These results demonstrate that untreated HIV-1 infected Moroccans will be susceptible to INSTIs.

The online version of this article (10.1186/s13104-018-3492-5) contains supplementary material, which is available to authorized users.

• HIV-1
• INSTIs
• Integrase
• Morocco
• Naïve patients
• Resistance mutations

• HIV-1 integrase
• LEDGF/p75
• inhibition
• interaction
• isoquinoline

The computational modeling of peptide inhibitors to target protein-protein binding interfaces is growing in interest as these are often too large, too shallow, and too feature-less for conventional small molecule compounds. Here, we present a rare successful application of an alchemical binding free energy method for the calculation of converged absolute binding free energies of a series of protein-peptide complexes. Specifically, we report the binding free energies of a series of cyclic peptides derived from the LEDGF/p75 protein to the integrase receptor of the HIV1 virus. The simulations recapitulate the effect of mutations relative to the wild-type binding motif of LEDGF/p75, providing structural, energetic and dynamical interpretations of the observed trends. The equilibration and convergence of the calculations are carefully analyzed. Convergence is aided by the adoption of a single-decoupling alchemical approach with implicit solvation, which circumvents the convergence difficulties of conventional double-decoupling protocols. We hereby present the single-decoupling methodology and critically evaluate its advantages and limitations. We also discuss some of the challenges and potential pitfalls of binding free energy calculations for complex molecular systems which have generally limited their applicability to the quantitative study of protein-peptide binding equilibria.

• HIV integrase
• LEDGF/p75
• alchemical calculations
• binding free energy
• implicit solvent
• protein-peptide complexes
• single-decoupling

• HIV integrase-binding domain
• domain swapping
• human immunodeficiency virus
• lens epithelium-derived growth factor

• Adaptor Proteins, Signal Transducing/chemistry
• Adaptor Proteins, Signal Transducing/isolation & purification
• Adaptor Proteins, Signal Transducing/metabolism
• Amino Acid Sequence
• Catalytic Domain
• Crystallization
• Crystallography, X-Ray
• HIV Integrase/chemistry
• HIV Integrase/metabolism
• Humans
• Models, Molecular
• Protein Conformation
• Transcription Factors/chemistry
• Transcription Factors/isolation & purification
• Transcription Factors/metabolism

• MR/K018779/1 Medical Research Council
• 2008-2011 Candlelighters
• Wellcome Trust
• 12051 Bloodwise
• MR/J000612/1 Medical Research Council
• 099246/Z/12/Z Wellcome Trust

• Cellular cofactors
• Protein complexes
• Protein–protein interaction
• Small organic molecules
• Viral replication
• Viruses
• Virus–host interactome
• Virus–host protein interactions

Preventing the protein-protein interaction of the cellular chromatin binding protein Lens Epithelium-Derived Growth Factor (LEDGF) and human immunodeficiency virus (HIV) integrase is an important possible strategy for anti-viral treatment for AIDS. We have used Intracellular Antibody Capture technology to isolate a single VH antibody domain that binds to LEDGF. The crystal structure of the LEDGF-VH complex reveals that the single domain antibody mimics the effect of binding of HIV integrase to LEDGF which is crucial for HIV propagation. CD4-expressing T cell lines were constructed to constitutively express the LEDGF-binding VH and these cells showed interference with HIV viral replication, assayed by virus capsid protein p24 production. Therefore, pre-conditioning cells to express antibody fragments confers effective intracellular immunization for preventing chronic viral replication and can be a way to prevent HIV spread in infected patients. This raises the prospect that intracellular immunization strategies that focus on cellular components of viral integrase protein interactions can be used to combat the problems associated with latent HIV virus re-emergence in patients. New genome editing development, such as using CRISPR/cas9, offer the prospect intracellularly immunized T cells in HIV+ patients.

• Anti-HIV
• Inhibitors
• Molecular docking
• Molecular dynamics
• Protein–protein interaction

Retroviral replication proceeds through the integration of a DNA copy of the viral RNA genome into the host cellular genome, a process that is mediated by the viral integrase (IN) protein. IN catalyzes two distinct chemical reactions: 3’-processing, whereby the viral DNA is recessed by a di- or trinucleotide at its 3’-ends, and strand transfer, in which the processed viral DNA ends are inserted into host chromosomal DNA. Although IN has been studied as a recombinant protein since the 1980s, detailed structural understanding of its catalytic functions awaited high resolution structures of functional IN-DNA complexes or intasomes, initially obtained in 2010 for the spumavirus prototype foamy virus (PFV). Since then, two additional retroviral intasome structures, from the α-retrovirus Rous sarcoma virus (RSV) and β-retrovirus mouse mammary tumor virus (MMTV), have emerged. Here, we briefly review the history of IN structural biology prior to the intasome era, and then compare the intasome structures of PFV, MMTV and RSV in detail. Whereas the PFV intasome is characterized by a tetrameric assembly of IN around the viral DNA ends, the newer structures harbor octameric IN assemblies. Although the higher order architectures of MMTV and RSV intasomes differ from that of the PFV intasome, they possess remarkably similar intasomal core structures. Thus, retroviral integration machineries have adapted evolutionarily to utilize disparate IN elements to construct convergent intasome core structures for catalytic function.

• DNA integration
• 3-dimensional structure
• Integrase
• Intasome
• Mouse mammary tumor virus
• Retrovirus
• Rous sarcoma virus
• Prototype foamy virus
• Human immunodeficiency virus/acquired immune deficiency syndrome

• R01 AI070042 NIAID NIH HHS

Integrase strand-transfer inhibitors (INSTIs), such as raltegravir (RAL), elvitegravir, or dolutegravir (DTG), are efficient antiretroviral agents used in HIV treatment in order to inhibit retroviral integration. By contrast to RAL treatments leading to well-identified mutation resistance pathways at the integrase level, recent clinical studies report several cases of patients failing DTG treatment without clearly identified resistance mutation in the integrase gene raising questions for the mechanism behind the resistance. These compounds, by impairing the integration of HIV-1 viral DNA into the host DNA, lead to an accumulation of unintegrated circular viral DNA forms. This viral DNA could be at the origin of the INSTI resistance by two different ways. The first one, sustained by a recent report, involves 2-long terminal repeat circles integration and the second one involves expression of accumulated unintegrated viral DNA leading to a basal production of viral particles maintaining the viral information.

• HIV-1
• integrase
• strand-transfer inhibitors
• unintegrated viral DNA
• viral resistance

• BiFC
• HIV-1
• HIV-1 integrase multimerization
• NCINIs

• Animals
• COS Cells
• Chlorocebus aethiops
• Cytological Techniques/methods
• HIV Integrase/metabolism
• HIV Integrase Inhibitors/analysis
• HIV-1/enzymology
• Optical Imaging/methods
• Protein Multimerization

• ZIA BC011105 Intramural NIH HHS

Transcription regulation in metazoans often involves promoter-proximal pausing of RNA polymerase (Pol) II, which requires the 4-subunit negative elongation factor (NELF). Here we discern the functional architecture of human NELF through X-ray crystallography, protein crosslinking, biochemical assays, and RNA crosslinking in cells. We identify a NELF core subcomplex formed by conserved regions in subunits NELF-A and NELF-C, and resolve its crystal structure. The NELF-AC subcomplex binds single-stranded nucleic acids in vitro, and NELF-C associates with RNA in vivo. A positively charged face of NELF-AC is involved in RNA binding, whereas the opposite face of the NELF-AC subcomplex binds NELF-B. NELF-B is predicted to form a HEAT repeat fold, also binds RNA in vivo, and anchors the subunit NELF-E, which is confirmed to bind RNA in vivo. These results reveal the three-dimensional architecture and three RNA-binding faces of NELF.

DOI:http://dx.doi.org/10.7554/eLife.14981.001

• RNA polymerase II
• biochemistry
• biophysics
• gene regulation
• gene transcription
• human
• promoter-proximal pausing
• structural biology

Persistence of latent, replication-competent Human Immunodeficiency Virus type 1 (HIV-1) provirus is the main impediment towards a cure for HIV/AIDS (Acquired Immune Deficiency Syndrome). Therefore, different therapeutic strategies to eliminate the viral reservoirs are currently being explored. We here propose a novel strategy to reduce the replicating HIV reservoir during primary HIV infection by means of drug-induced retargeting of HIV integration. A novel class of integration inhibitors, referred to as LEDGINs, inhibit the interaction between HIV integrase and the LEDGF/p75 host cofactor, the main determinant of lentiviral integration site selection. We show for the first time that LEDGF/p75 depletion hampers HIV-1 reactivation in cell culture. Next we demonstrate that LEDGINs relocate and retarget HIV integration resulting in a HIV reservoir that is refractory to reactivation by different latency-reversing agents. Taken together, these results support the potential of integrase inhibitors that modulate integration site targeting to reduce the likeliness of viral rebound.

•

LEDGF/p75 depletion hampers HIV reactivation in cell culture.

Different strategies to cure HIV infection are being explored. Although complete eradication of the HIV provirus is the ultimate goal, disease remission allowing treatment interruption without viral rebound would constitute a significant leap forward. HIV integration site selection is orchestrated by LEDGF/p75. The advent of LEDGINs, that block the interaction between integrase and LEDGF/p75, allowed us to examine the hypothesis that interference with HIV integration site selection would yield integration sites that are less optimal for productive infection. Here we provide evidence in cell culture that LEDGIN treatment during acute HIV infection yields an HIV reservoir refractory to reactivation.

• HIV latency
• HIV remission
• Integration
• LEDGF/p75
• LEDGIN

Drug design against human immunodeficiency virus type 1 (HIV-1) integrase through its mechanistic study is of great interest in the area in biological research. The main obstacle in this area is the absence of the full-length crystal structure for HIV-1 integrase to be used as a model. A complete structure, similar to HIV-1 of a prototype foamy virus integrase in complex with DNA, including all conservative residues, is available and has been extensively used in recent investigations.

The aim of this study was to determine whether the above model is precisely representative of HIV-1 integrase. This would critically determine the success of any designed drug using the model in deactivation of integrase and AIDS treatment.

Primarily, a new structure for HIV-1 was constructed, using a crystal structure of prototype foamy virus as the starting structure. The constructed structure of HIV-1 integrase was simultaneously simulated with a prototype foamy virus integrase on a separate occasion.

Our results indicate that the HIV-1 system behaves differently from the prototype foamy virus in terms of folding, hydration, hydrophobicity of binding site and stability.

Based on our findings, we can conclude that HIV-1 integrase is vastly different from the prototype foamy virus integrase and does not resemble it, and the modeling output of the prototype foamy virus simulations could not be simply generalized to HIV-1 integrase. Therefore, our HIV-1 model seems to be more representative and more useful for future research.

• Human Immunodeficiency Virus Type-1
• Integrase
• Molecular Dynamic Simulation
• Prototype Foamy Virus

• BET family transcription factor
• ET binding motif (EBM)
• NMR
• protein–protein interaction
• retroviral integration

• HIV-1
• integrase
• strand-transfer inhibitors
• unintegrated viral DNA
• viral resistance