scholarly journals Characterization of 67 Confirmed Clustered Regularly Interspaced Short Palindromic Repeats Loci in 52 Strains of Staphylococci

2021 ◽  
Vol 12 ◽  
Author(s):  
Ying Wang ◽  
Tingting Mao ◽  
Yinxia Li ◽  
Wenwei Xiao ◽  
Xuan Liang ◽  
...  
Keyword(s):  
Structural Characteristics ◽  
Direct Repeat ◽  
Minimum Free Energy ◽  
Important Species ◽  
Crispr Locus ◽  
Adaptive Immune ◽  
Associated Proteins ◽  
The Relationship ◽  
Cas Proteins

Staphylococcus aureus (S. aureus), which is one of the most important species of Staphylococci, poses a great threat to public health. Clustered regularly interspaced short palindromic repeats (CRISPR) and their CRISPR-associated proteins (Cas) are an adaptive immune platform to combat foreign mobile genetic elements (MGEs) such as plasmids and phages. The aim of this study is to describe the distribution and structure of CRISPR-Cas system in S. aureus, and to explore the relationship between CRISPR and horizontal gene transfer (HGT). Here, we analyzed 67 confirmed CRISPR loci and 15 companion Cas proteins in 52 strains of Staphylococci with bioinformatics methods. Comparing with the orphan CRISPR loci in Staphylococci, the strains harboring complete CRISPR-Cas systems contained multiple CRISPR loci, direct repeat sequences (DR) forming stable RNA secondary structures with lower minimum free energy (MFE), and variable spacers with detectable protospacers. In S. aureus, unlike the orphan CRISPRs away from Staphylococcal cassette chromosome mec (SCCmec), the complete CRISPR-Cas systems were in J1 region of SCCmec. In addition, we found a conserved motif 5′-TTCTCGT-3′ that may protect their downstream sequences from DNA interference. In general, orphan CRISPR locus in S. aureus differed greatly from the structural characteristics of the CRISPR-Cas system. Collectively, our results provided new insight into the diversity and characterization of the CRISPR-Cas system in S. aureus.

scholarly journals CRISPR-Cas: biology, mechanisms and relevance

2016 ◽  
Vol 371 (1707) ◽  
pp. 20150496 ◽  
Author(s):  
Frank Hille ◽  
Emmanuelle Charpentier
Keyword(s):  
Dna Sequences ◽  
Adaptive Immune System ◽  
Crispr Locus ◽  
Biochemical Processes ◽  
Adaptive Immune ◽  
Physiological Properties ◽  
Target Dna ◽  
Associated Proteins ◽  
Cas Proteins

Prokaryotes have evolved several defence mechanisms to protect themselves from viral predators. Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) display a prokaryotic adaptive immune system that memorizes previous infections by integrating short sequences of invading genomes—termed spacers—into the CRISPR locus. The spacers interspaced with repeats are expressed as small guide CRISPR RNAs (crRNAs) that are employed by Cas proteins to target invaders sequence-specifically upon a reoccurring infection. The ability of the minimal CRISPR-Cas9 system to target DNA sequences using programmable RNAs has opened new avenues in genome editing in a broad range of cells and organisms with high potential in therapeutical applications. While numerous scientific studies have shed light on the biochemical processes behind CRISPR-Cas systems, several aspects of the immunity steps, however, still lack sufficient understanding. This review summarizes major discoveries in the CRISPR-Cas field, discusses the role of CRISPR-Cas in prokaryotic immunity and other physiological properties, and describes applications of the system as a DNA editing technology and antimicrobial agent. This article is part of the themed issue ‘The new bacteriology’.

Fuel characteristics and fire behaviour in mature Mediterranean gorse shrublands

2004 ◽  
Vol 13 (1) ◽  
pp. 79 ◽  
Author(s):  
Martín De Luis ◽  
Manuel J. Baeza ◽  
José Raventós ◽  
José C. González-Hidalgo
Keyword(s):  
Forest Fires ◽  
Plant Density ◽  
Structural Characteristics ◽  
Fire Severity ◽  
Fire Behaviour ◽  
Fuel Distribution ◽  
Degree Of Dominance ◽  
Mediterranean Regions ◽  
The Relationship

Since the early 1990s, Mediterranean gorse shrublands have expanded significantly in the Mediterranean regions of Spain mainly as a result of the increase in the frequency and extension of forest fires. Mediterranean gorse (Ulex parviflorus), which has been described as a degradation stage of forest communities after fire, has also been described as a fire-prone community. Thus, its presence increases the risk that new fires might occur. In spite of this evidence, there is little information on both the composition and structural characteristics of these communities or the relationship that might exist between these vegetation characteristics and fire behaviour. In this paper we present the results of a characterization of the vegetative structure (plant density, specific composition, biomass fractions, and horizontal and vertical fuel distribution) in Mediterranean gorse. We also analyse fire behaviour using indicators obtained at different scales. Our results show mature Mediterranean gorse shrublands to be communities with high biomass values (3000–4000 g m−2) and high horizontal and vertical vegetation continuity, in which the proportion of fine dead fuel fractions with low moisture content is around 50% of the total phytomass present. Ulex parviflorus is the dominant species and its degree of dominance is a key element in the behaviour of fire. Both the fire-line intensity values and the fire severity values observed can be considered high with respect to those observed in other Mediterranean communities, thus confirming Mediterranean gorse as a high-risk community.

scholarly journals Cas13b is a Type VI-B CRISPR-associated RNA-Guided RNase differentially regulated by accessory proteins Csx27 and Csx28

2016 ◽  
Author(s):  
Aaron Smargon ◽  
David B.T. Cox ◽  
Neena Pyzocha ◽  
Kaijie Zheng ◽  
Ian M. Slaymaker ◽  
...  
Keyword(s):  
Rna Interference ◽  
Essential Gene ◽  
Sequence Database ◽  
Flanking Sequence ◽  
E Coli ◽  
Adaptive Immune ◽  
Associated Proteins ◽  
Adaptive Immune Systems ◽  
Target Rna

CRISPR-Cas adaptive immune systems defend microbes against foreign nucleic acids via RNA-guided endonucleases. Using a computational sequence database mining approach, we identify two Class 2 CRISPR-Cas systems (subtype VI-B) that lack Cas1 and Cas2 and encompass a single large effector protein, Cas13b, along with one of two previously uncharacterized associated proteins, Csx27 or Csx28. We establish that these CRISPR-Cas systems can achieve RNA interference when heterologously expressed. Through a combination of biochemical and genetic experiments, we show that Cas13b processes its own CRISPR array with short and long direct repeats, cleaves target RNA, and exhibits collateral RNase activity. Using an E. coli essential gene screen, we demonstrate that Cas13b has a double-sided protospacer-flanking sequence and elucidate RNA secondary structure requirements for targeting. We also find that Csx27 represses, whereas Csx28 enhances, Cas13b-mediated RNA interference. Characterization of these CRISPR systems creates opportunities to develop tools to manipulate and monitor cellular transcripts.

Structural Characterization of Micron-scaled Reticulated Copper Foams

2011 ◽  
Vol 1304 ◽  
Author(s):  
Stephanie J Lin ◽  
Jason H. Nadler
Keyword(s):  
Image Analysis ◽  
Structural Characteristics ◽  
Porous Network ◽  
Unique Challenge ◽  
Copper Foam ◽  
Fluid Permeability ◽  
Capillary Fluid ◽  
Relationship Of ◽  
The Relationship

AbstractThe development of a multifunctional, micron-scaled, reticulated copper foam that reliably exhibits high intrinsic thermal conductivity, efficient capillary fluid and evaporative transport over a wide area presents a unique challenge. In this work, the relationship of critical foam processing variables such as sintering temperature and template size on the pore size distribution and pore neck/body ratio is investigated using image analysis. The resulting fluid permeability values of these foams are estimated by using the Kozeny Carman equation and the porosity, surface area per unit area and tortuosity obtained through image analysis. Estimating the fluid permeability of these foams is useful for predicting the mass and heat transfer within the porous network, and provides a metric for optimizing the foam’s structural characteristics for a particular application.

scholarly journals Type I-E CRISPR-Cas system as an immune system in a eukaryote

2018 ◽  
Author(s):  
Devashish Rath ◽  
Lina Amlinger ◽  
Gargi Bindal ◽  
Magnus Lundgren
Keyword(s):  
Nucleic Acid ◽  
Dna Cleavage ◽  
Basic Research ◽  
Type I ◽  
Crispr Locus ◽  
Adaptive Immune ◽  
Research Platform ◽  
Adaptive Immune Systems ◽  
Cas Proteins

AbstractDefense against viruses and other mobile genetic elements (MGEs) is important in many organisms. The CRISPR-Cas systems found in bacteria and archaea constitute adaptive immune systems that acquire the ability to recognize MGEs by introducing nucleic acid samples, spacers, in the CRISPR locus. The CRISPR is transcribed and processed, and the produced CRISPR RNAs guide Cas proteins to degrade matching nucleic acid sequences. No CRISPR-Cas system is found to occur naturally in eukaryotic cells but here we demonstrate interference by type I-E CRISPR-Cas system from Escherichia coli introduced in Saccharomyces cerevisiae. The designed CRISPR arrays are properly expressed and processed in S. cerevisiae. Targeted plasmids display reduced transformation efficiency, indicative of DNA cleavage. Unlike e.g. Cas9-based systems, which can be used to inactivate MGEs in eukaryotes by introducing specific mutations, type I-E systems processively degrade the target. The type I-E system thus allows for defense without knowledge of MGE gene function. The reconstituted CRISPR-Cas system in S. cerevisiae can also function as a basic research platform for testing the role of various factors in the interference process.

scholarly journals Assembly and translocation of a CRISPR-Cas primed acquisition complex

2017 ◽  
Author(s):  
Maxwell W. Brown ◽  
Kaylee E. Dillard ◽  
Yibei Xiao ◽  
Adam Dolan ◽  
Erik Hernandez ◽  
...  
Keyword(s):  
Single Molecule ◽  
Target Recognition ◽  
Adaptive Immune Response ◽  
Type I ◽  
Crispr Locus ◽  
One Dimensional ◽  
Adaptive Immune ◽  
Dna Loop ◽  
Recognition Efficiency

AbstractBacteria and archaea destroy foreign nucleic acids by mounting an RNA-based CRISPR-Cas adaptive immune response1–3. In type I CRISPR-Cas systems, the most frequently found type of CRISPR in bacteria and archaea3,4, foreign DNAs that trigger efficient immunity can also provoke primed acquisition of protospacers into the CRISPR locus5–12. Both interference and primed acquisition require Cascade (CRISPR-associated complex for antiviral defense) and the Cas3 helicase/nuclease. Primed acquisition also requires the Cas1-Cas2 integrase; however, the biophysical mechanisms of how interference and primed acquisition are coordinated have remained elusive. Here, we present single-molecule characterization of the type I-E Thermobifida fusca (Tfu) primed acquisition complex (PAC). TfuCascade rapidly samples non-specific DNA for its target via facilitated one-dimensional (1D) diffusion. An evolutionary-conserved positive patch on the Cse1 subunit increases the target recognition efficiency by promoting this 1D diffusion. Cas3 loads at target-bound Cascade and the Cascade/Cas3 complex initiates processive translocation via a looped DNA intermediate. Moving Cascade/Cas3 complexes stall and release the DNA loop at protein roadblocks. Cas1-Cas2 samples DNA transiently via 3D collisions, but stably associates with target-bound Cascade. Cas1-Cas2 also remains associated with translocating Cascade/Cas3, forming the PAC. By directly imaging all key subcomplexes involved in target recognition, interference, and primed acquisition, this work provides a molecular basis for the coordinated steps in CRISPR-based adaptive immunity.

Molecular Mechanisms of CRISPR-Cas Immunity in Bacteria

2020 ◽  
Vol 54 (1) ◽  
pp. 93-120 ◽  
Author(s):  
Philip M. Nussenzweig ◽  
Luciano A. Marraffini
Keyword(s):  
Immune Response ◽  
Molecular Mechanisms ◽  
Adaptive Immune System ◽  
Crispr Locus ◽  
Defense Strategies ◽  
Adaptive Immune ◽  
Third Phase ◽  
Short Palindromic Repeat ◽  
The Many ◽  
Cas Proteins

Prokaryotes have developed numerous defense strategies to combat the constant threat posed by the diverse genetic parasites that endanger them. Clustered regularly interspaced short palindromic repeat (CRISPR)-Cas loci guard their hosts with an adaptive immune system against foreign nucleic acids. Protection starts with an immunization phase, in which short pieces of the invader's genome, known as spacers, are captured and integrated into the CRISPR locus after infection. Next, during the targeting phase, spacers are transcribed into CRISPR RNAs (crRNAs) that guide CRISPR-associated (Cas) nucleases to destroy the invader's DNA or RNA. Here we describe the many different molecular mechanisms of CRISPR targeting and how they are interconnected with the immunization phase through a third phase of the CRISPR-Cas immune response: primed spacer acquisition. In this phase, Cas proteins direct the crRNA-guided acquisition of additional spacers to achieve a more rapid and robust immunization of the population.

Skeletal elements of crinoid echinoderms: High-resolution structural and microanalytical results

1983 ◽  
Vol 41 ◽  
pp. 194-195
Author(s):  
D. F. Blake ◽  
L. F. Allard ◽  
D. R. Peacor
Keyword(s):  
High Resolution ◽  
Marine Invertebrates ◽  
Sea Urchins ◽  
Structural Features ◽  
Sea Stars ◽  
High Resolution Tem ◽  
Relationship Of ◽  
The Relationship ◽  
Insight Into

Echinodermata is a phylum of marine invertebrates which has been extant since Cambrian time (c.a. 500 m.y. before the present). Modern examples of echinoderms include sea urchins, sea stars, and sea lilies (crinoids). The endoskeletons of echinoderms are composed of plates or ossicles (Fig. 1) which are with few exceptions, porous, single crystals of high-magnesian calcite. Despite their single crystal nature, fracture surfaces do not exhibit the near-perfect {10.4} cleavage characteristic of inorganic calcite. This paradoxical mix of biogenic and inorganic features has prompted much recent work on echinoderm skeletal crystallography. Furthermore, fossil echinoderm hard parts comprise a volumetrically significant portion of some marine limestones sequences. The ultrastructural and microchemical characterization of modern skeletal material should lend insight into: 1). The nature of the biogenic processes involved, for example, the relationship of Mg heterogeneity to morphological and structural features in modern echinoderm material, and 2). The nature of the diagenetic changes undergone by their ancient, fossilized counterparts. In this study, high resolution TEM (HRTEM), high voltage TEM (HVTEM), and STEM microanalysis are used to characterize tha ultrastructural and microchemical composition of skeletal elements of the modern crinoid Neocrinus blakei.

Microstructural Characterization of Au-Ge-Ni based ohmic contacts to GaAs/AlGaAs modfet device

1987 ◽  
Vol 45 ◽  
pp. 326-327
Author(s):  
A.K. Rai ◽  
A.K. Petford-Long ◽  
A. Ezis ◽  
D.W. Langer
Keyword(s):  
Contact Resistance ◽  
Ohmic Contact ◽  
Ohmic Contacts ◽  
Microstructural Characterization ◽  
Almost Everywhere ◽  
Spacer Layer ◽  
Good Contact ◽  
The Relationship ◽  
Lower Contact

Considerable amount of work has been done in studying the relationship between the contact resistance and the microstructure of the Au-Ge-Ni based ohmic contacts to n-GaAs. It has been found that the lower contact resistivity is due to the presence of Ge rich and Au free regions (good contact area) in contact with GaAs. Thus in order to obtain an ohmic contact with lower contact resistance one should obtain a uniformly alloyed region of good contact areas almost everywhere. This can possibly be accomplished by utilizing various alloying schemes. In this work microstructural characterization, employing TEM techniques, of the sequentially deposited Au-Ge-Ni based ohmic contact to the MODFET device is presented.The substrate used in the present work consists of 1 μm thick buffer layer of GaAs grown on a semi-insulating GaAs substrate followed by a 25 Å spacer layer of undoped AlGaAs.

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