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Nature Reviews. Microbiology Jan 2022A hallmark of retroviral replication is establishment of the proviral state, wherein a DNA copy of the viral RNA genome is stably incorporated into a host cell... (Review)
Review
A hallmark of retroviral replication is establishment of the proviral state, wherein a DNA copy of the viral RNA genome is stably incorporated into a host cell chromosome. Integrase is the viral enzyme responsible for the catalytic steps involved in this process, and integrase strand transfer inhibitors are widely used to treat people living with HIV. Over the past decade, a series of X-ray crystallography and cryogenic electron microscopy studies have revealed the structural basis of retroviral DNA integration. A variable number of integrase molecules congregate on viral DNA ends to assemble a conserved intasome core machine that facilitates integration. The structures additionally informed on the modes of integrase inhibitor action and the means by which HIV acquires drug resistance. Recent years have witnessed the development of allosteric integrase inhibitors, a highly promising class of small molecules that antagonize viral morphogenesis. In this Review, we explore recent insights into the organization and mechanism of the retroviral integration machinery and highlight open questions as well as new directions in the field.
Topics: Crystallography, X-Ray; DNA, Viral; HIV Integrase; HIV-1; Humans; Integrases; Models, Molecular; Protein Conformation; Retroviridae; Virus Integration
PubMed: 34244677
DOI: 10.1038/s41579-021-00586-9 -
Nature Jun 2023CRISPR-Cas adaptive immune systems capture DNA fragments from invading mobile genetic elements and integrate them into the host genome to provide a template for...
CRISPR-Cas adaptive immune systems capture DNA fragments from invading mobile genetic elements and integrate them into the host genome to provide a template for RNA-guided immunity. CRISPR systems maintain genome integrity and avoid autoimmunity by distinguishing between self and non-self, a process for which the CRISPR/Cas1-Cas2 integrase is necessary but not sufficient. In some microorganisms, the Cas4 endonuclease assists CRISPR adaptation, but many CRISPR-Cas systems lack Cas4. Here we show here that an elegant alternative pathway in a type I-E system uses an internal DnaQ-like exonuclease (DEDDh) to select and process DNA for integration using the protospacer adjacent motif (PAM). The natural Cas1-Cas2/exonuclease fusion (trimmer-integrase) catalyses coordinated DNA capture, trimming and integration. Five cryo-electron microscopy structures of the CRISPR trimmer-integrase, visualized both before and during DNA integration, show how asymmetric processing generates size-defined, PAM-containing substrates. Before genome integration, the PAM sequence is released by Cas1 and cleaved by the exonuclease, marking inserted DNA as self and preventing aberrant CRISPR targeting of the host. Together, these data support a model in which CRISPR systems lacking Cas4 use fused or recruited exonucleases for faithful acquisition of new CRISPR immune sequences.
Topics: CRISPR-Associated Proteins; CRISPR-Cas Systems; Cryoelectron Microscopy; DNA; Exonucleases; Integrases; Biocatalysis; Genome, Bacterial
PubMed: 37316664
DOI: 10.1038/s41586-023-06178-2 -
Current Opinion in Structural Biology Dec 2017Retroviral DNA integration takes place in the context of the intasome nucleoprotein complex. X-ray crystal structures of functional spumaviral intasomes were previously... (Review)
Review
Retroviral DNA integration takes place in the context of the intasome nucleoprotein complex. X-ray crystal structures of functional spumaviral intasomes were previously revealed to harbor a homotetramer of integrase, and it was generally believed that integrase tetramers catalyzed the integration of other retroviruses. The elucidation of new structures from four different retroviruses over the past year has however revealed this is not the case. The number of integrase molecules required to construct the conserved intasome core structure differs between viral species. While four subunits suffice for spumaviruses, α- and β-retroviruses require eight and the lentiviruses use up to sixteen. Herein we described these alternative architectures, highlighting both evolutionary and structural constraints that result in the different integrase-DNA stoichiometries across Retroviridae.
Topics: DNA, Viral; Integrases; Models, Molecular; Multiprotein Complexes; Nucleic Acid Conformation; Protein Binding; Protein Conformation; Retroviridae; Structure-Activity Relationship; Virus Integration
PubMed: 28458055
DOI: 10.1016/j.sbi.2017.04.005 -
Annual Review of Virology Nov 2015The retroviral integrases are virally encoded, specialized recombinases that catalyze the insertion of viral DNA into the host cell's DNA, a process that is essential... (Review)
Review
The retroviral integrases are virally encoded, specialized recombinases that catalyze the insertion of viral DNA into the host cell's DNA, a process that is essential for virus propagation. We have learned a great deal since the existence of an integrated form of retroviral DNA (the provirus) was first proposed by Howard Temin in 1964. Initial studies focused on the genetics and biochemistry of avian and murine virus DNA integration, but the pace of discovery increased substantially with advances in technology, and an influx of investigators focused on the human immunodeficiency virus. We begin with a brief account of the scientific landscape in which some of the earliest discoveries were made, and summarize research that led to our current understanding of the biochemistry of integration. A more detailed account of recent analyses of integrase structure follows, as they have provided valuable insights into enzyme function and raised important new questions.
Topics: Animals; Humans; Integrases; Models, Molecular; Retroviridae; Retroviridae Infections; Viral Proteins; Virus Integration
PubMed: 26958915
DOI: 10.1146/annurev-virology-100114-055043 -
Current Opinion in Microbiology Aug 2017DNA site-specific recombinases are enzymes (often associated with mobile DNA elements) that catalyse breaking and rejoining of DNA strands at specific points, thereby... (Review)
Review
DNA site-specific recombinases are enzymes (often associated with mobile DNA elements) that catalyse breaking and rejoining of DNA strands at specific points, thereby bringing about precise genetic rearrangements. Serine integrases are a group of recombinases derived from bacteriophages. Their unusual properties, including directionality of recombination and simple site requirements, are leading to their development as efficient, versatile tools for applications in experimental biology, biotechnology, synthetic biology and gene therapy. This article summarizes our current knowledge of serine integrase structure and mechanism, then outlines key factors that affect the performance of these phage recombination systems. Recently published studies, that have expanded the repertoire of available systems and reveal system-specific characteristics, will help us to choose the best integrases for envisaged applications.
Topics: Bacteriophages; Biotechnology; Gene Targeting; Genetic Therapy; Integrases; Models, Biological; Molecular Biology; Recombination, Genetic
PubMed: 28599144
DOI: 10.1016/j.mib.2017.04.006 -
Current Opinion in Pediatrics Dec 2022To provide an update on the current understanding of the role of wingless/integrase-1 (Wnt) signaling in pediatric allergic asthma and other pediatric lung diseases. (Review)
Review
PURPOSE OF REVIEW
To provide an update on the current understanding of the role of wingless/integrase-1 (Wnt) signaling in pediatric allergic asthma and other pediatric lung diseases.
RECENT FINDINGS
The Wnt signaling pathway is critical for normal lung development. Genetic and epigenetic human studies indicate a link between Wnt signaling and the development and severity of asthma in children. Mechanistic studies using animal models of allergic asthma demonstrate a key role for Wnt signaling in allergic airway inflammation and remodeling. More recently, data on bronchopulmonary dysplasia (BPD) pathogenesis points to the Wnt signaling pathway as an important regulator.
SUMMARY
Current data indicates that the Wnt signaling pathway is an important mediator in allergic asthma and BPD pathogenesis. Further studies are needed to characterize the roles of individual Wnt signals in childhood disease, and to identify potential novel therapeutic targets to slow or prevent disease processes.
Topics: Infant, Newborn; Animals; Humans; Child; Integrases; Asthma; Lung; Wnt Signaling Pathway; Bronchopulmonary Dysplasia
PubMed: 36081387
DOI: 10.1097/MOP.0000000000001173 -
Cells Mar 2022Integrons are powerful recombination systems found in bacteria, which act as platforms capable of capturing, stockpiling, excising and reordering mobile elements called... (Review)
Review
Integrons are powerful recombination systems found in bacteria, which act as platforms capable of capturing, stockpiling, excising and reordering mobile elements called cassettes. These dynamic genetic machineries confer a very high potential of adaptation to their host and have quickly found themselves at the forefront of antibiotic resistance, allowing for the quick emergence of multi-resistant phenotypes in a wide range of bacterial species. Part of the success of the integron is explained by its ability to integrate various environmental and biological signals in order to allow the host to respond to these optimally. In this review, we highlight the substantial interconnectivity that exists between integrons and their hosts and its importance to face changing environments. We list the factors influencing the expression of the cassettes, the expression of the integrase, and the various recombination reactions catalyzed by the integrase. The combination of all these host factors allows for a very tight regulation of the system at the cost of a limited ability to spread by horizontal gene transfer and function in remotely related hosts. Hence, we underline the important consequences these factors have on the evolution of integrons. Indeed, we propose that sedentary chromosomal integrons that were less connected or connected via more universal factors are those that have been more successful upon mobilization in mobile genetic structures, in contrast to those that were connected to species-specific host factors. Thus, the level of specificity of the involved host factors network may have been decisive for the transition from chromosomal integrons to the mobile integrons, which are now widespread. As such, integrons represent a perfect example of the conflicting relationship between the ability to control a biological system and its potential for transferability.
Topics: Bacteria; Drug Resistance, Microbial; Gene Transfer, Horizontal; Integrases; Integrons
PubMed: 35326376
DOI: 10.3390/cells11060925 -
ACS Synthetic Biology Feb 2018Serine integrases catalyze precise rearrangement of DNA through site-specific recombination of small sequences of DNA called attachment (att) sites. Unlike other... (Review)
Review
Serine integrases catalyze precise rearrangement of DNA through site-specific recombination of small sequences of DNA called attachment (att) sites. Unlike other site-specific recombinases, the recombination reaction driven by serine integrases is highly directional and can only be reversed in the presence of an accessory protein called a recombination directionality factor (RDF). The ability to control reaction directionality has led to the development of serine integrases as tools for controlled rearrangement and modification of DNA in synthetic biology, gene therapy, and biotechnology. This review discusses recent advances in serine integrase technologies focusing on their applications in genome engineering, DNA assembly, and logic and data storage devices.
Topics: Attachment Sites, Microbiological; DNA; Integrases; Recombination, Genetic; Synthetic Biology
PubMed: 29316791
DOI: 10.1021/acssynbio.7b00308 -
Viruses Nov 2022Integrase Strand Transfer Inhibitors (INSTIs) are currently used as the most effective therapy in the treatment of human immunodeficiency virus (HIV) infections.... (Review)
Review
Integrase Strand Transfer Inhibitors (INSTIs) are currently used as the most effective therapy in the treatment of human immunodeficiency virus (HIV) infections. Raltegravir (RAL) and Elvitegravir (EVG), the first generation of INSTIs used successfully in clinical treatment, are susceptible to the emergence of viral resistance and have a high rate of cross-resistance. To counteract these resistant mutants, second-generation INSTI drugs have been developed: Dolutegravir (DTG), Cabotegravir (CAB), and Bictegravir (BIC). However, HIV is also able to develop resistance mechanisms against the second-generation of INSTIs. This review describes the mode of action of INSTIs and then summarizes and evaluates some typical resistance mutations, such as substitution and insertion mutations. The role of unintegrated viral DNA is also discussed as a new pathway involved in conferring resistance to INSTIs. This allows us to have a more detailed understanding of HIV resistance to these inhibitors, which may contribute to the development of new INSTIs in the future.
Topics: Humans; HIV Integrase Inhibitors; Raltegravir Potassium; HIV Infections; Heterocyclic Compounds, 3-Ring; Mutation; Integrases; HIV Integrase; Drug Resistance, Viral
PubMed: 36560595
DOI: 10.3390/v14122591 -
Microbiology Spectrum Dec 2014Due to the importance of human immunodeficiency virus type 1 (HIV-1) integrase as a drug target, the biochemistry and structural aspects of retroviral DNA integration... (Review)
Review
Due to the importance of human immunodeficiency virus type 1 (HIV-1) integrase as a drug target, the biochemistry and structural aspects of retroviral DNA integration have been the focus of intensive research during the past three decades. The retroviral integrase enzyme acts on the linear double-stranded viral DNA product of reverse transcription. Integrase cleaves specific phosphodiester bonds near the viral DNA ends during the 3' processing reaction. The enzyme then uses the resulting viral DNA 3'-OH groups during strand transfer to cut chromosomal target DNA, which simultaneously joins both viral DNA ends to target DNA 5'-phosphates. Both reactions proceed via direct transesterification of scissile phosphodiester bonds by attacking nucleophiles: a water molecule for 3' processing, and the viral DNA 3'-OH for strand transfer. X-ray crystal structures of prototype foamy virus integrase-DNA complexes revealed the architectures of the key nucleoprotein complexes that form sequentially during the integration process and explained the roles of active site metal ions in catalysis. X-ray crystallography furthermore elucidated the mechanism of action of HIV-1 integrase strand transfer inhibitors, which are currently used to treat AIDS patients, and provided valuable insights into the mechanisms of viral drug resistance.
Topics: Catalytic Domain; Crystallography, X-Ray; DNA; DNA, Viral; Integrases; Metals; Protein Conformation; Recombination, Genetic; Retroviridae
PubMed: 26104441
DOI: 10.1128/microbiolspec.MDNA3-0024-2014