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Journal of Molecular Biology Aug 2019The β-lactams retain a central place in the antibacterial armamentarium. In Gram-negative bacteria, β-lactamase enzymes that hydrolyze the amide bond of the... (Review)
Review
The β-lactams retain a central place in the antibacterial armamentarium. In Gram-negative bacteria, β-lactamase enzymes that hydrolyze the amide bond of the four-membered β-lactam ring are the primary resistance mechanism, with multiple enzymes disseminating on mobile genetic elements across opportunistic pathogens such as Enterobacteriaceae (e.g., Escherichia coli) and non-fermenting organisms (e.g., Pseudomonas aeruginosa). β-Lactamases divide into four classes; the active-site serine β-lactamases (classes A, C and D) and the zinc-dependent or metallo-β-lactamases (MBLs; class B). Here we review recent advances in mechanistic understanding of each class, focusing upon how growing numbers of crystal structures, in particular for β-lactam complexes, and methods such as neutron diffraction and molecular simulations, have improved understanding of the biochemistry of β-lactam breakdown. A second focus is β-lactamase interactions with carbapenems, as carbapenem-resistant bacteria are of grave clinical concern and carbapenem-hydrolyzing enzymes such as KPC (class A) NDM (class B) and OXA-48 (class D) are proliferating worldwide. An overview is provided of the changing landscape of β-lactamase inhibitors, exemplified by the introduction to the clinic of combinations of β-lactams with diazabicyclooctanone and cyclic boronate serine β-lactamase inhibitors, and of progress and strategies toward clinically useful MBL inhibitors. Despite the long history of β-lactamase research, we contend that issues including continuing unresolved questions around mechanism; opportunities afforded by new technologies such as serial femtosecond crystallography; the need for new inhibitors, particularly for MBLs; the likely impact of new β-lactam:inhibitor combinations and the continuing clinical importance of β-lactams mean that this remains a rewarding research area.
Topics: Anti-Bacterial Agents; Carbapenem-Resistant Enterobacteriaceae; Carbapenems; Catalytic Domain; Drug Combinations; Drug Resistance, Bacterial; Enterobacteriaceae; Gram-Negative Bacteria; Humans; Interspersed Repetitive Sequences; beta-Lactamase Inhibitors; beta-Lactamases; beta-Lactams
PubMed: 30959050
DOI: 10.1016/j.jmb.2019.04.002 -
Microbiology Spectrum Jul 2019Enterococci are unusually well adapted for survival and persistence in a variety of adverse environments, including on inanimate surfaces in the hospital environment and... (Review)
Review
Enterococci are unusually well adapted for survival and persistence in a variety of adverse environments, including on inanimate surfaces in the hospital environment and at sites of infection. This intrinsic ruggedness undoubtedly played a role in providing opportunities for enterococci to interact with other overtly drug-resistant microbes and acquire additional resistances on mobile elements. The rapid rise of antimicrobial resistance among hospital-adapted enterococci has rendered hospital-acquired infections a leading therapeutic challenge. With about a quarter of a genome of additional DNA conveyed by mobile elements, there are undoubtedly many more properties that have been acquired that help enterococci persist and spread in the hospital setting and cause diseases that have yet to be defined. Much remains to be learned about these ancient and rugged microbes, particularly in the area of pathogenic mechanisms involved with human diseases.
Topics: Animals; Enterococcus; Gram-Positive Bacterial Infections; Humans; Interspersed Repetitive Sequences; Virulence
PubMed: 31298205
DOI: 10.1128/microbiolspec.GPP3-0053-2018 -
Nature Chemical Biology Jan 2021Many bacterial and archaeal organisms use clustered regularly interspaced short palindromic repeats-CRISPR associated (CRISPR-Cas) systems to defend themselves from... (Review)
Review
Many bacterial and archaeal organisms use clustered regularly interspaced short palindromic repeats-CRISPR associated (CRISPR-Cas) systems to defend themselves from mobile genetic elements. These CRISPR-Cas systems are classified into six types based on their composition and mechanism. CRISPR-Cas enzymes are widely used for genome editing and offer immense therapeutic opportunity to treat genetic diseases. To realize their full potential, it is important to control the timing, duration, efficiency and specificity of CRISPR-Cas enzyme activities. In this Review we discuss the mechanisms of natural CRISPR-Cas regulatory biomolecules and engineering strategies that enhance or inhibit CRISPR-Cas immunity by altering enzyme function. We also discuss the potential applications of these CRISPR regulators and highlight unanswered questions about their evolution and purpose in nature.
Topics: Antibiosis; Archaea; Bacteria; Bacteriophages; CRISPR-Associated Protein 9; CRISPR-Cas Systems; Clustered Regularly Interspaced Short Palindromic Repeats; Gene Editing; Gene Expression Regulation, Archaeal; Gene Expression Regulation, Bacterial; Genetic Engineering; Humans; Interspersed Repetitive Sequences; RNA, Guide, CRISPR-Cas Systems
PubMed: 33328654
DOI: 10.1038/s41589-020-00700-7 -
Nature Reviews. Molecular Cell Biology Jul 2022Transposable elements (TEs) comprise about half of the mammalian genome. TEs often contain sequences capable of recruiting the host transcription machinery, which they... (Review)
Review
Transposable elements (TEs) comprise about half of the mammalian genome. TEs often contain sequences capable of recruiting the host transcription machinery, which they use to express their own products and promote transposition. However, the regulatory sequences carried by TEs may affect host transcription long after the TEs have lost the ability to transpose. Recent advances in genome analysis and engineering have facilitated systematic interrogation of the regulatory activities of TEs. In this Review, we discuss diverse mechanisms by which TEs contribute to transcription regulation. Notably, TEs can donate enhancer and promoter sequences that influence the expression of host genes, modify 3D chromatin architecture and give rise to novel regulatory genes, including non-coding RNAs and transcription factors. We discuss how TEs spur regulatory evolution and facilitate the emergence of genetic novelties in mammalian physiology and development. By virtue of their repetitive and interspersed nature, TEs offer unique opportunities to dissect the effects of mutation and genomic context on the function and evolution of cis-regulatory elements. We argue that TE-centric studies hold the key to unlocking general principles of transcription regulation and evolution.
Topics: Animals; DNA Transposable Elements; Evolution, Molecular; Gene Expression Regulation; Mammals; Promoter Regions, Genetic; Regulatory Sequences, Nucleic Acid; Transcription Factors
PubMed: 35228718
DOI: 10.1038/s41580-022-00457-y -
Cell Jun 2023Retroelements are the widespread jumping elements considered as major drivers for genome evolution, which can also be repurposed as gene-editing tools. Here, we...
Retroelements are the widespread jumping elements considered as major drivers for genome evolution, which can also be repurposed as gene-editing tools. Here, we determine the cryo-EM structures of eukaryotic R2 retrotransposon with ribosomal DNA target and regulatory RNAs. Combined with biochemical and sequencing analysis, we reveal two essential DNA regions, Drr and Dcr, required for recognition and cleavage. The association of 3' regulatory RNA with R2 protein accelerates the first-strand cleavage, blocks the second-strand cleavage, and initiates the reverse transcription starting from the 3'-tail. Removing 3' regulatory RNA by reverse transcription allows the association of 5' regulatory RNA and initiates the second-strand cleavage. Taken together, our work explains the DNA recognition and RNA supervised sequential retrotransposition mechanisms by R2 machinery, providing insights into the retrotransposon and application reprogramming.
Topics: Retroelements; RNA; DNA Cleavage; RNA-Directed DNA Polymerase; Reverse Transcription
PubMed: 37301196
DOI: 10.1016/j.cell.2023.05.032 -
Science (New York, N.Y.) Apr 2021Long-read and strand-specific sequencing technologies together facilitate the de novo assembly of high-quality haplotype-resolved human genomes without parent-child trio...
Long-read and strand-specific sequencing technologies together facilitate the de novo assembly of high-quality haplotype-resolved human genomes without parent-child trio data. We present 64 assembled haplotypes from 32 diverse human genomes. These highly contiguous haplotype assemblies (average minimum contig length needed to cover 50% of the genome: 26 million base pairs) integrate all forms of genetic variation, even across complex loci. We identified 107,590 structural variants (SVs), of which 68% were not discovered with short-read sequencing, and 278 SV hotspots (spanning megabases of gene-rich sequence). We characterized 130 of the most active mobile element source elements and found that 63% of all SVs arise through homology-mediated mechanisms. This resource enables reliable graph-based genotyping from short reads of up to 50,340 SVs, resulting in the identification of 1526 expression quantitative trait loci as well as SV candidates for adaptive selection within the human population.
Topics: Female; Genetic Variation; Genome, Human; Genotype; Haplotypes; High-Throughput Nucleotide Sequencing; Humans; INDEL Mutation; Interspersed Repetitive Sequences; Male; Population Groups; Quantitative Trait Loci; Retroelements; Sequence Analysis, DNA; Sequence Inversion; Whole Genome Sequencing
PubMed: 33632895
DOI: 10.1126/science.abf7117 -
Cell Dec 2022The perinatal period represents a critical window for cognitive and immune system development, promoted by maternal and infant gut microbiomes and their metabolites....
The perinatal period represents a critical window for cognitive and immune system development, promoted by maternal and infant gut microbiomes and their metabolites. Here, we tracked the co-development of microbiomes and metabolomes from late pregnancy to 1 year of age using longitudinal multi-omics data from a cohort of 70 mother-infant dyads. We discovered large-scale mother-to-infant interspecies transfer of mobile genetic elements, frequently involving genes associated with diet-related adaptations. Infant gut metabolomes were less diverse than maternal but featured hundreds of unique metabolites and microbe-metabolite associations not detected in mothers. Metabolomes and serum cytokine signatures of infants who received regular-but not extensively hydrolyzed-formula were distinct from those of exclusively breastfed infants. Taken together, our integrative analysis expands the concept of vertical transmission of the gut microbiome and provides original insights into the development of maternal and infant microbiomes and metabolomes during late pregnancy and early life.
Topics: Female; Humans; Infant; Pregnancy; Gastrointestinal Microbiome; Microbiota; Mothers; Breast Feeding; Feces; Interspersed Repetitive Sequences
PubMed: 36563663
DOI: 10.1016/j.cell.2022.11.023 -
Cell Aug 2022Non-allelic recombination between homologous repetitive elements contributes to evolution and human genetic disorders. Here, we combine short- and long-DNA read...
Non-allelic recombination between homologous repetitive elements contributes to evolution and human genetic disorders. Here, we combine short- and long-DNA read sequencing of repeat elements with a new bioinformatics pipeline to show that somatic recombination of Alu and L1 elements is widespread in the human genome. Our analysis uncovers tissue-specific non-allelic homologous recombination hallmarks; moreover, we find that centromeres and cancer-associated genes are enriched for retroelements that may act as recombination hotspots. We compare recombination profiles in human-induced pluripotent stem cells and differentiated neurons and find that the neuron-specific recombination of repeat elements accompanies chromatin changes during cell-fate determination. Finally, we report that somatic recombination profiles are altered in Parkinson's and Alzheimer's disease, suggesting a link between retroelement recombination and genomic instability in neurodegeneration. This work highlights a significant contribution of the somatic recombination of repeat elements to genomic diversity in health and disease.
Topics: Alu Elements; Genome, Human; Homologous Recombination; Humans; Long Interspersed Nucleotide Elements; Repetitive Sequences, Nucleic Acid; Retroelements
PubMed: 35882231
DOI: 10.1016/j.cell.2022.06.032 -
Plasmid Sep 2019
Topics: Humans; Interspersed Repetitive Sequences; Plasmids
PubMed: 31472205
DOI: 10.1016/j.plasmid.2019.102437 -
Nature Jul 2023Whole-genome synthesis provides a powerful approach for understanding and expanding organism function. To build large genomes rapidly, scalably and in parallel, we need...
Whole-genome synthesis provides a powerful approach for understanding and expanding organism function. To build large genomes rapidly, scalably and in parallel, we need (1) methods for assembling megabases of DNA from shorter precursors and (2) strategies for rapidly and scalably replacing the genomic DNA of organisms with synthetic DNA. Here we develop bacterial artificial chromosome (BAC) stepwise insertion synthesis (BASIS)-a method for megabase-scale assembly of DNA in Escherichia coli episomes. We used BASIS to assemble 1.1 Mb of human DNA containing numerous exons, introns, repetitive sequences, G-quadruplexes, and long and short interspersed nuclear elements (LINEs and SINEs). BASIS provides a powerful platform for building synthetic genomes for diverse organisms. We also developed continuous genome synthesis (CGS)-a method for continuously replacing sequential 100 kb stretches of the E. coli genome with synthetic DNA; CGS minimizes crossovers between the synthetic DNA and the genome such that the output for each 100 kb replacement provides, without sequencing, the input for the next 100 kb replacement. Using CGS, we synthesized a 0.5 Mb section of the E. coli genome-a key intermediate in its total synthesis-from five episomes in 10 days. By parallelizing CGS and combining it with rapid oligonucleotide synthesis and episome assembly, along with rapid methods for compiling a single genome from strains bearing distinct synthetic genome sections, we anticipate that it will be possible to synthesize entire E. coli genomes from functional designs in less than 2 months.
Topics: Humans; DNA; Escherichia coli; Genome, Bacterial; Plasmids; Repetitive Sequences, Nucleic Acid; Synthetic Biology; Chromosomes, Artificial, Bacterial; Exons; Introns; G-Quadruplexes; Long Interspersed Nucleotide Elements; Short Interspersed Nucleotide Elements; Oligodeoxyribonucleotides; Time Factors
PubMed: 37380776
DOI: 10.1038/s41586-023-06268-1