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DNA Repair Sep 2020To ensure genome integrity, the joining of breaks in the phosphodiester backbone of duplex DNA is required during DNA replication and to complete the repair of almost... (Review)
Review
To ensure genome integrity, the joining of breaks in the phosphodiester backbone of duplex DNA is required during DNA replication and to complete the repair of almost all types of DNA damage. In human cells, this task is accomplished by DNA ligases encoded by three genes, LIG1, LIG3 and LIG4. Mutations in LIG1 and LIG4 have been identified as the causative factor in two inherited immunodeficiency syndromes. Moreover, there is emerging evidence that DNA ligases may be good targets for the development of novel anti-cancer agents. In this graphical review, we provide an overview of the roles of the DNA ligases encoded by the three human LIG genes in DNA replication and repair.
Topics: DNA; DNA Damage; DNA Ligase ATP; DNA Repair; DNA Replication; Humans; Poly-ADP-Ribose Binding Proteins
PubMed: 33087274
DOI: 10.1016/j.dnarep.2020.102908 -
Trends in Biochemical Sciences Jul 2022Mutations in BRCA1 and BARD1 predispose carriers to breast and ovarian cancers. The BRCA1 and BARD1 proteins form a heterodimeric complex (BRCA1/BARD1) that regulates... (Review)
Review
Mutations in BRCA1 and BARD1 predispose carriers to breast and ovarian cancers. The BRCA1 and BARD1 proteins form a heterodimeric complex (BRCA1/BARD1) that regulates many biological processes, including transcription and DNA double-stranded break repair. These functions are mediated by the only known enzymatic activity of BRCA1/BARD1 in its capacity as an E3 ubiquitin ligase and its role as a central hub for many large protein complexes. But the mechanisms by which BRCA1/BARD1 interfaces with chromatin, where it exerts its major functions, have remained unknown. Here, we review recent advancements in structural and cellular biology that have provided critical insights into how BRCA1/BARD1 serves as both a nucleosome reader and writer to facilitate transcriptional regulation and DNA repair by homologous recombination.
Topics: BRCA1 Protein; DNA Breaks, Double-Stranded; DNA Repair; Nucleosomes; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases
PubMed: 35351360
DOI: 10.1016/j.tibs.2022.03.001 -
Frontiers in Microbiology 2023DNA ligase is an important enzyme ubiquitous in all three kingdoms of life that can ligate DNA strands, thus playing essential roles in DNA replication, repair and... (Review)
Review
DNA ligase is an important enzyme ubiquitous in all three kingdoms of life that can ligate DNA strands, thus playing essential roles in DNA replication, repair and recombination . , DNA ligase is also used in biotechnological applications requiring in DNA manipulation, including molecular cloning, mutation detection, DNA assembly, DNA sequencing, and other aspects. Thermophilic and thermostable enzymes from hyperthermophiles that thrive in the high-temperature (above 80°C) environments have provided an important pool of useful enzymes as biotechnological reagents. Similar to other organisms, each hyperthermophile harbors at least one DNA ligase. In this review, we summarize recent progress on structural and biochemical properties of thermostable DNA ligases from hyperthermophiles, focusing on similarities and differences between DNA ligases from hyperthermophilic bacteria and archaea, and between these thermostable DNA ligases and non-thermostable homologs. Additionally, altered thermostable DNA ligases are discussed. Possessing improved fidelity or thermostability compared to the wild-type enzymes, they could be potential DNA ligases for biotechnology in the future. Importantly, we also describe current applications of thermostable DNA ligases from hyperthermophiles in biotechnology.
PubMed: 37293226
DOI: 10.3389/fmicb.2023.1198784 -
Molecular Microbiology Oct 2022Adenoviruses (AdVs) are widespread in vertebrates. They infect the respiratory and gastrointestinal tracts, the eyes, heart, liver, and kidney, and are lethal to... (Review)
Review
Adenoviruses (AdVs) are widespread in vertebrates. They infect the respiratory and gastrointestinal tracts, the eyes, heart, liver, and kidney, and are lethal to immunosuppressed people. Mastadenoviruses infecting mammals comprise several hundred different types, and many specifically infect humans. Human adenoviruses are the most widely used vectors in clinical applications, including cancer treatment and COVID-19 vaccination. AdV vectors are physically and genetically stable and generally safe in humans. The particles have an icosahedral coat and a nucleoprotein core with a DNA genome. We describe the concept of AdV cell entry and highlight recent advances in cytoplasmic transport, uncoating, and nuclear import of the viral DNA. We highlight a recently discovered "linchpin" function of the virion protein V ensuring cytoplasmic particle stability, which is relaxed at the nuclear pore complex by cues from the E3 ubiquitin ligase Mind bomb 1 (MIB1) and the proteasome triggering disruption. Capsid disruption by kinesin motor proteins and microtubules exposes the linchpin and renders protein V a target for MIB1 ubiquitination, which dissociates V from viral DNA and enhances DNA nuclear import. These advances uncover mechanisms controlling capsid stability and premature uncoating and provide insight into nuclear transport of nucleic acids.
Topics: Animals; Humans; Active Transport, Cell Nucleus; Adenoviridae; DNA, Viral; Proteasome Endopeptidase Complex; Kinesins; COVID-19 Vaccines; COVID-19; Nuclear Pore; Capsid Proteins; Ubiquitin-Protein Ligases; Nucleoproteins; Mammals
PubMed: 35434852
DOI: 10.1111/mmi.14909 -
Cell Host & Microbe Oct 2021The pore-forming protein gasdermin D (GSDMD) executes lytic cell death called pyroptosis to eliminate the replicative niche of intracellular pathogens. Evolution favors...
The pore-forming protein gasdermin D (GSDMD) executes lytic cell death called pyroptosis to eliminate the replicative niche of intracellular pathogens. Evolution favors pathogens that circumvent this host defense mechanism. Here, we show that the Shigella ubiquitin ligase IpaH7.8 functions as an inhibitor of GSDMD. Shigella is an enteroinvasive bacterium that causes hemorrhagic gastroenteritis in primates, but not rodents. IpaH7.8 contributes to species specificity by ubiquitinating human, but not mouse, GSDMD and targeting it for proteasomal degradation. Accordingly, infection of human epithelial cells with IpaH7.8-deficient Shigella flexneri results in increased GSDMD-dependent cell death compared with wild type. Consistent with pyroptosis contributing to murine disease resistance, eliminating GSDMD from NLRC4-deficient mice, which are already sensitized to oral infection with Shigella flexneri, leads to further enhanced bacterial replication and increased disease severity. This work highlights a species-specific pathogen arms race focused on maintenance of host cell viability.
Topics: Animals; Bacterial Proteins; Dysentery, Bacillary; Epithelial Cells; Female; Host-Pathogen Interactions; Humans; Mice; Mice, Knockout; Phosphate-Binding Proteins; Pore Forming Cytotoxic Proteins; Proteolysis; Shigella flexneri; Ubiquitin-Protein Ligases
PubMed: 34492225
DOI: 10.1016/j.chom.2021.08.010 -
The Journal of Clinical Investigation Jan 2024As the leading cause of disability worldwide, low back pain (LBP) is recognized as a pivotal socioeconomic challenge to the aging population and is largely attributed to...
As the leading cause of disability worldwide, low back pain (LBP) is recognized as a pivotal socioeconomic challenge to the aging population and is largely attributed to intervertebral disc degeneration (IVDD). Elastic nucleus pulposus (NP) tissue is essential for the maintenance of IVD structural and functional integrity. The accumulation of senescent NP cells with an inflammatory hypersecretory phenotype due to aging and other damaging factors is a distinctive hallmark of IVDD initiation and progression. In this study, we reveal a mechanism of IVDD progression in which aberrant genomic DNA damage promoted NP cell inflammatory senescence via activation of the cyclic GMP-AMP synthase/stimulator of IFN genes (cGAS/STING) axis but not of absent in melanoma 2 (AIM2) inflammasome assembly. Ataxia-telangiectasia-mutated and Rad3-related protein (ATR) deficiency destroyed genomic integrity and led to cytosolic mislocalization of genomic DNA, which acted as a powerful driver of cGAS/STING axis-dependent inflammatory phenotype acquisition during NP cell senescence. Mechanistically, disassembly of the ATR-tripartite motif-containing 56 (ATR-TRIM56) complex with the enzymatic liberation of ubiquitin-specific peptidase 5 (USP5) and TRIM25 drove changes in ATR ubiquitination, with ATR switching from K63- to K48-linked modification, c thereby promoting ubiquitin-proteasome-dependent dynamic instability of ATR protein during NP cell senescence progression. Importantly, an engineered extracellular vesicle-based strategy for delivering ATR-overexpressing plasmid cargo efficiently diminished DNA damage-associated NP cell senescence and substantially mitigated IVDD progression, indicating promising targets and effective approaches to ameliorate the chronic pain and disabling effects of IVDD.
Topics: Humans; Aged; Intervertebral Disc Degeneration; Nucleus Pulposus; Aging; Cellular Senescence; Nucleotidyltransferases; Intervertebral Disc; Tripartite Motif Proteins; Ubiquitin-Protein Ligases; Ataxia Telangiectasia Mutated Proteins
PubMed: 38488012
DOI: 10.1172/JCI165140 -
Essays in Biochemistry Aug 2023Viruses have developed sophisticated biochemical and genetic mechanisms to manipulate and exploit their hosts. Enzymes derived from viruses have been essential research... (Review)
Review
Viruses have developed sophisticated biochemical and genetic mechanisms to manipulate and exploit their hosts. Enzymes derived from viruses have been essential research tools since the first days of molecular biology. However, most viral enzymes that have been commercialized are derived from a small number of cultivated viruses, which is remarkable considering the extraordinary diversity and abundance of viruses revealed by metagenomic analysis. Given the explosion of new enzymatic reagents derived from thermophilic prokaryotes over the past 40 years, those obtained from thermophilic viruses should be equally potent tools. This review discusses the still-limited state of the art regarding the functional biology and biotechnology of thermophilic viruses with a focus on DNA polymerases, ligases, endolysins, and coat proteins. Functional analysis of DNA polymerases and primase-polymerases from phages infecting Thermus, Aquificaceae, and Nitratiruptor has revealed new clades of enzymes with strong proofreading and reverse transcriptase capabilities. Thermophilic RNA ligase 1 homologs have been characterized from Rhodothermus and Thermus phages, with both commercialized for circularization of single-stranded templates. Endolysins from phages infecting Thermus, Meiothermus, and Geobacillus have shown high stability and unusually broad lytic activity against Gram-negative and Gram-positive bacteria, making them targets for commercialization as antimicrobials. Coat proteins from thermophilic viruses infecting Sulfolobales and Thermus strains have been characterized, with diverse potential applications as molecular shuttles. To gauge the scale of untapped resources for these proteins, we also document over 20,000 genes encoded by uncultivated viral genomes from high-temperature environments that encode DNA polymerase, ligase, endolysin, or coat protein domains.
Topics: Viruses; Bacteriophages; Bacteria; DNA-Directed DNA Polymerase; Biotechnology; Ligases; Biology
PubMed: 37222046
DOI: 10.1042/EBC20220209 -
Biochemical Society Transactions Jun 2020DNA replication is a complex process that needs to be executed accurately before cell division in order to maintain genome integrity. DNA replication is divided into... (Review)
Review
DNA replication is a complex process that needs to be executed accurately before cell division in order to maintain genome integrity. DNA replication is divided into three main stages: initiation, elongation and termination. One of the key events during initiation is the assembly of the replicative helicase at origins of replication, and this mechanism has been very well described over the last decades. In the last six years however, researchers have also focused on deciphering the molecular mechanisms underlying the disassembly of the replicative helicase during termination. Similar to replisome assembly, the mechanism of replisome disassembly is strictly regulated and well conserved throughout evolution, although its complexity increases in higher eukaryotes. While budding yeast rely on just one pathway for replisome disassembly in S phase, higher eukaryotes evolved an additional mitotic pathway over and above the default S phase specific pathway. Moreover, replisome disassembly has been recently found to be a key event prior to the repair of certain DNA lesions, such as under-replicated DNA in mitosis and inter-strand cross-links (ICLs) in S phase. Although replisome disassembly in human cells has not been characterised yet, they possess all of the factors involved in these pathways in model organisms, and de-regulation of many of them are known to contribute to tumorigenesis and other pathological conditions.
Topics: Animals; Caenorhabditis elegans; Cell Cycle; Cell Cycle Proteins; Cell Survival; DNA; DNA Replication; Genome; Humans; Minichromosome Maintenance Complex Component 7; Mitosis; Neoplasms; Protein Interaction Mapping; Replication Origin; S Phase; Saccharomyces cerevisiae; Saccharomycetales; Ubiquitin; Ubiquitin-Protein Ligases; Ubiquitination; Xenopus laevis
PubMed: 32490508
DOI: 10.1042/BST20190363 -
Autoimmunity and immunodeficiency associated with monoallelic LIG4 mutations via haploinsufficiency.The Journal of Allergy and Clinical... Aug 2023Biallelic mutations in LIG4 encoding DNA-ligase 4 cause a rare immunodeficiency syndrome manifesting as infant-onset life-threatening and/or opportunistic infections,...
BACKGROUND
Biallelic mutations in LIG4 encoding DNA-ligase 4 cause a rare immunodeficiency syndrome manifesting as infant-onset life-threatening and/or opportunistic infections, skeletal malformations, radiosensitivity and neoplasia. LIG4 is pivotal during DNA repair and during V(D)J recombination as it performs the final DNA-break sealing step.
OBJECTIVES
This study explored whether monoallelic LIG4 missense mutations may underlie immunodeficiency and autoimmunity with autosomal dominant inheritance.
METHODS
Extensive flow-cytometric immune-phenotyping was performed. Rare variants of immune system genes were analyzed by whole exome sequencing. DNA repair functionality and T-cell-intrinsic DNA damage tolerance was tested with an ensemble of in vitro and in silico tools. Antigen-receptor diversity and autoimmune features were characterized by high-throughput sequencing and autoantibody arrays. Reconstitution of wild-type versus mutant LIG4 were performed in LIG4 knockout Jurkat T cells, and DNA damage tolerance was subsequently assessed.
RESULTS
A novel heterozygous LIG4 loss-of-function mutation (p.R580Q), associated with a dominantly inherited familial immune-dysregulation consisting of autoimmune cytopenias, and in the index patient with lymphoproliferation, agammaglobulinemia, and adaptive immune cell infiltration into nonlymphoid organs. Immunophenotyping revealed reduced naive CD4 T cells and low TCR-Vα7.2 T cells, while T-/B-cell receptor repertoires showed only mild alterations. Cohort screening identified 2 other nonrelated patients with the monoallelic LIG4 mutation p.A842D recapitulating clinical and immune-phenotypic dysregulations observed in the index family and displaying T-cell-intrinsic DNA damage intolerance. Reconstitution experiments and molecular dynamics simulations categorize both missense mutations as loss-of-function and haploinsufficient.
CONCLUSIONS
This study provides evidence that certain monoallelic LIG4 mutations may cause human immune dysregulation via haploinsufficiency.
Topics: Humans; DNA Ligases; Autoimmunity; Haploinsufficiency; DNA Ligase ATP; Immunologic Deficiency Syndromes; Mutation; DNA
PubMed: 37004747
DOI: 10.1016/j.jaci.2023.03.022 -
International Journal of Molecular... Aug 2021UBR box E3 ligases, also called N-recognins, are integral components of the N-degron pathway. Representative N-recognins include UBR1, UBR2, UBR4, and UBR5, and they... (Review)
Review
UBR box E3 ligases, also called N-recognins, are integral components of the N-degron pathway. Representative N-recognins include UBR1, UBR2, UBR4, and UBR5, and they bind destabilizing N-terminal residues, termed N-degrons. Understanding the molecular bases of their substrate recognition and the biological impact of the clearance of their substrates on cellular signaling pathways can provide valuable insights into the regulation of these pathways. This review provides an overview of the current knowledge of the binding mechanism of UBR box N-recognin/N-degron interactions and their roles in signaling pathways linked to G-protein-coupled receptors, apoptosis, mitochondrial quality control, inflammation, and DNA damage. The targeting of these UBR box N-recognins can provide potential therapies to treat diseases such as cancer and neurodegenerative diseases.
Topics: Animals; Apoptosis; DNA Damage; Humans; Inflammation; Receptors, G-Protein-Coupled; Signal Transduction; Ubiquitin-Protein Ligases
PubMed: 34361089
DOI: 10.3390/ijms22158323